Dnase co-expression in host cells

Abstract

The present invention relates to the introduction of expression constructs encoding different gene products such as proteins or nucleic acids at defined ratios into host cell lines containing multiple dock sites for insertion of the nucleic acid construct, and to the production of proteins that require expression of at least two gene products.

Description

DNASE CO-EXPRESSION IN HOST CELLSCROSS-REFERENCE TO RELATED APPLICATIONSThe present application claims priority to U.S. Provisional Application No. 63 / 536,279, filed September 1, 2023, which is incorporated herein by reference in its entirety.SEQUENCE LISTINGThe text of the computer readable sequence listing filed herewith, titled “CATA_41307_601_SequenceListing.xml”, created August 28, 2024, having a file size of 13,008 bytes, is hereby incorporated by reference in its entirety.FIELD OF THE INVENTIONThe present invention relates to the introduction of expression constructs encoding different gene products such as proteins or nucleic acids at defined ratios into host cell lines containing multiple dock sites for insertion of the nucleic acid construct, and to the production of proteins that require expression of at least two gene products.BACKGROUND OF THE INVENTIONTherapeutic recombinant products are extensively used for human medical purposes. Recombinant proteins can be expressed in various types of living organisms. CHO and HEK cells are some of the most commonly used host cells for the manufacture of biopharmaceuticals. Although using the host cells for the production of therapeutics has many advantages, contamination of biopharmaceuticals with host cell DNA is a major concern for manufacturers. Genomic DNA contamination in biopharmaceuticals has been considered as a possible risk factor for patients receiving recombinant protein pharmaceuticals.Thus, there is a need for biopharmaceutical manufacturers to monitor DNA contamination and keep the quantity of impurities in bioproducts as low as a safety limit suggested by the regulatory authorities such as FDA, which generally must be lower than 100 pg of DNA per milligram of protein. Removal of unwanted DNA can be expensive, time consuming, and reduce the efficiency of recovery of the biopharmaceutical protein.Accordingly, what is needed in the art are approaches for limiting the contamination of biopharmaccutical protein preparations by host cell DNA.SUMMARY OF THE INVENTIONThe present invention relates to the introduction of expression constructs encoding different gene products such as proteins or nucleic acids at defined ratios into host cell lines containing multiple dock sites for insertion of the nucleic acid construct, and to the production of proteins that require expression of at least two gene products. For example, the present invention provides host cells that allow for co-expression DNase with a product of interest to improve expression and / or purification of the protein of interest from the host cell culture. As another example, the present invention also provides host cells and associated methods that allow for expression of two or more products of interest at varying ratios in a host cell.Accordingly, in some preferred embodiments, the present invention provides an eukaryotic host cell comprising a first exogenous nucleic acid sequence encoding DNase 1 operably linked to a promoter sequence and at least a second exogenous nucleic acid sequence encoding a first product of interest operably linked to a promoter sequence, wherein the first sequence encoding DNase 1 and the at least second exogenous encoding the first product of interest are co-expressed. In some preferred embodiments, the host cell further comprises a third exogenous nucleic acid sequence encoding a second product of interest. In some preferred embodiments, the host cell further comprises a fourth exogenous nucleic acid sequence encoding a third product of interest. In some preferred embodiments, the host cell further comprises a fifth exogenous nucleic acid sequence encoding a fourth product of interest. In some preferred embodiments, the host cell further comprises a sixth exogenous nucleic acid sequence encoding a fifth product of interest. In some preferred embodiments, the sequence encoding DNase 1 is further operably linked to a secretion signal sequence.In some preferred embodiments, the one or more products of interest is a protein or proteins. In some preferred embodiments, the first product of interest is selected from the group consisting of an immunoglobulin heavy chain sequence and an immunoglobulin light chain sequence. In some preferred embodiments, the host cell further comprises a third exogenous nucleic acid sequence encoding a second product of interest operably linked to a promoter sequence and a secretion signal sequence, wherein the first product of interest is animmunoglobulin light chain sequence and the second product of interest is an immunoglobulin heavy chain sequence.In some preferred embodiments, the one or more products of interest are a nucleic acid or nucleic acids. In some preferred embodiments, the at least a second exogenous nucleic acid is a viral nucleic acid. In some preferred embodiments, the viral nucleic acids are selected from the group consisting of retroviral nucleic acids, lentiviral nucleic acids, adenoviral nucleic acids, and adeno-associated virus (AAV) nucleic acids. In some preferred embodiments, the at least a second exogenous nucleic acid is an exosomal nucleic acid.In some preferred embodiments, the first exogenous nucleic acid sequence and at least a second exogenous nucleic acid sequence are stably integrated into the genome of the host cell.In some preferred embodiments, from 2 to 500 copies each of the first exogenous nucleic acid sequence and at least a second exogenous nucleic acid sequence are stably integrated into the genome of the host cell. In some preferred embodiments, from 5 to 200 copies each of the first exogenous nucleic acid sequence and at least a second exogenous nucleic acid sequence are stably integrated into the genome of the host cell.In some preferred embodiments, the ratio of the integrated first exogenous nucleic acid sequence to the integrated second exogenous nucleic acid sequence is from 1:1 to 1:500, 1:1 to 1:400, 1:1 to 1:300, 1:1 to 1:300, 1:1 to 1:200, or 1:1 to 1:100. In some preferred embodiments, the ratio of the integrated first exogenous nucleic acid sequence to the integrated second exogenous nucleic acid sequence is from 1:2 to 1:500, 1:2 to 1:400, 1:2 to 1:300, 1:2 to 1:300, 1:2 to 1:200, or 1:2 to 1:100. In some preferred embodiments, the ratio of the integrated first exogenous nucleic acid sequence to the integrated second exogenous nucleic acid sequence is from 1:3 to 1:500, 1:3 to 1:400, 1:3 to 1:300, 1:3 to 1:300, 1:3 to 1:200, or 1:3 to 1:100. In some preferred embodiments, the ratio of the integrated first exogenous nucleic acid sequence to the integrated second exogenous nucleic acid sequence is from 1:5 to 1:500, 1:5 to 1:400, 1:5 to 1:300, 1:5 to 1:300, 1:5 to 1:200, or 1:5 to 1:100. In some preferred embodiments, the ratio of the integrated first exogenous nucleic acid sequence to the integrated second exogenous nucleic acid sequence is from 1:10 to 1:500, 1:10 to 1:400, 1:10 to 1:300, 1:10 to 1:300, 1:10 to 1:200, or 1:10 to 1:100.In some preferred embodiments, the first exogenous nucleic acid sequence and at least a second exogenous nucleic acid sequence are stably integrated into the genome of the host cell ata docking site. In some preferred embodiments, the docking site comprises at least one dock site insertion element and the exogenous nucleic acid sequences each comprise at least one insertion element compatible with the at least one dock site insertion element in the integrated docking sites.In some preferred embodiments, the exogenous nucleic acid sequences further comprise at least the following elements in operable association in 5’ to 3’ order: an internal promoter sequence; a nucleic acid sequence encoding the first product of interest or second product that is operably linked to the internal promoter; and a poly A signal sequence.In some preferred embodiments, the exogenous nucleic acid sequences comprise a selectable marker sequence. In some preferred embodiments, the exogenous nucleic acid sequences comprise different selectable marker sequences. In some preferred embodiments, one of the exogenous nucleic acid sequences comprises a selectable marker sequence and the other of the exogenous nucleic acid sequences does not comprise a selectable marker sequence. In some preferred embodiments, the selectable marker sequences are 5’ to the internal promoter sequence and are operably linked to a 5’ promoter sequence.In some preferred embodiments, the exogenous nucleic acid sequences comprise an extending packaging region (EPR) between the 5 ’ promoter and the selectable marker. In some preferred embodiments, the EPR comprises multiple potential Kozak sequences and / or ATG translation start sites.In some preferred embodiments, the promoter sequence is selected from the group consisting of SIN-LTR, SV40, EFla, E. coli lac, E. coli trp, phage lambda PL, phage lambda PR, T3, T7, cytomegalovirus (CMV) immediate early, herpes simplex virus (HSV) thymidine kinase, alpha-lactalbumin, and mouse metallothionein-I promoter sequences. In some preferred embodiments, the first promoter sequence is a weak promoter sequence. In some preferred embodiments, the first promoter sequence is not a retroviral LTR promoter.In some preferred embodiments, the integrated docking sites further comprise an exogenous promoter. In some preferred embodiments, the exogenous promoter is selected from the group consisting of SIN-LTR, SV40, EFl , E. coli lac, E. coli trp, phage lambda PL, phage lambda PR, T3, T7, cytomegalovirus (CMV) immediate early, herpes simplex virus (HSV) thymidine kinase, alpha-lactalbumin, and mouse metallothionein-I promoter sequences. In somepreferred embodiments, the promoter is a retroviral LTR. In some preferred embodiments, the retroviral LTR is a SIN LTR.In some preferred embodiments, the host cell line comprises a nucleic acid construct encoding an enzyme that facilitates insertion of the nucleic acid expression construct at the dock site. In some preferred embodiments, the nucleic acid construct encoding an enzyme that facilitates insertion of the exogenous nucleic acid sequences at the dock site is provided in a vector. In some preferred embodiments, the vector is a plasmid vector. In some preferred embodiments, the enzyme is selected from the group consisting of an integrase, a recombinase, a nuclease and a nickase.In some preferred embodiments, the host cell genome comprises from 5 to 500 integrated docking sites, each docking site comprising at least one dock site insertion element. In some preferred embodiments, the integrated docking sites are independently positioned throughout the host cell genome. In some preferred embodiments, the dock site insertion element is targeted by enzyme selected from the group consisting of an integrase, a recombinase, a nuclease and a nickase.In some preferred embodiments, the dock site insertion element is selected from the group consisting of a recombinase dock site insertion element and a HDR dock site insertion element. In some preferred embodiments, the dock site insertion element is a recombinase dock site insertion element. In some preferred embodiments, the recombinase dock site insertion element comprises an attachment site (att). In some preferred embodiments, the attachment site (att) is selected from the group consisting of attB and attP and attR and attL. In some preferred embodiments, the recombinase dock site insertion element comprises a LoxP sequence. In some preferred embodiments, the recombinase dock site insertion element is a Flp Recombination Target (FRT) site. In some preferred embodiments, the dock site insertion element is a HDR dock site insertion element. In some preferred embodiments, the HDR dock site insertion element comprises one or two dock site homology arms. In some preferred embodiments, the HDR dock site insertion element further comprises one or more sequences homologous to a guide RNA sequence. In some preferred embodiments, the dock site homology arms are from about 30 to 1000 bases in length. In some preferred embodiments, each docking site is flanked by exogenous integrating vector sequences. In some pref emed embodiments, the exogenous integrating vector sequences are selected from the group consisting of viral vector sequences andtransposon vector sequences. In some preferred embodiments, the docking sites each further comprise a sequence encoding a selectable maker operably linked to a promoter.In some preferred embodiments, the dock site insertion element is positioned to facilitate cassette exchange. In some preferred embodiments, each docking site comprises two dock site insertion elements. In some preferred embodiments, the two dock site insertion elements are positioned to facilitate cassette exchange. In some preferred embodiments, the two dock site insertion elements flank sequences encoding a selectable marker, an enzyme, or a combination thereof. In some preferred embodiments, the exogenous nucleic acid sequences further comprise a signal peptide sequence operably linked to the first product of interest.In some preferred embodiments, the host cell is selected from the group consisting of Chinese Hamster Ovary (CHO) cells, HEK 293 cells, CAP cells, bovine mammary epithelial cells, monkey kidney CV1 line transformed by SV40, baby hamster kidney cells, mouse sertoli cells, monkey kidney cells, African green monkey kidney cells, human cervical carcinoma cells, canine kidney cells, buffalo rat liver cells, human lung cells, human liver cells, mouse mammary tumor, TRI cells, MRC 5 cells, FS4 cells, rat fibroblasts, MDBK cells and human hepatoma line cells. In some preferred embodiments, the host cell is selected from the group consisting of a Chinese Hamster Ovary (CHO) cells, a HEK 293 cells and a CAP cells. In some preferred embodiments, the host cell line is a GS knockout cell line. In some preferred embodiments, the host cell line is a DHFR knockout cell line.In some preferred embodiments, the present invention provides a cell culture comprising host cells as described above.In some preferred embodiments, the present invention provides methods comprising : culturing a plurality of host cells as described above in a culture medium under conditions such that the product of interest or second products of interests and / or an assembled complex thereof is produced; and purifying the product or products of interest or assembled complex thereof. In some preferred embodiments, the product or products of interest are secreted into the culture medium and purified therefrom.In another aspect, the present invention relates to the introduction of expression constructs encoding different gene products such as proteins or nucleic acids at defined ratios into host cell lines containing multiple dock sites for insertion of the nucleic acid construct, and to the production of proteins that require expression of at least two gene products.In some preferred embodiments, the present invention provides methods comprising introducing at least first nucleic acid constructs encoding a first protein of interest or nucleic acid of interest and second nucleic acid constructs encoding a second protein of interest or nucleic acid of interest at a ratio of first nucleic acid constructs to second nucleic acid constructs of from 1:1 to 5000:1 (including ranges therein, e.g., 1:1 to 4000:1, 1:1: 3000:1, 1:1 to 2000:1, 1:1 to 1000:1, etc.) into a host cell having genome comprising from 1 to 500 integrated docking sites, each docking site comprising at least one dock site insertion element and the nucleic acid constructs each comprising at least one insertion element compatible with the at least one dock site insertion element in the integrated docking sites, under conditions such that the nucleic acid expression constructs are inserted at the dock sites at a ratio of first nucleic acid constructs to second nucleic acid constructs of at least 1:1. In some preferred embodiments, the ratio of inserted first nucleic acid constructs to second nucleic acid constructs is from 1.1:1 to 1000:1, 500:1, 200:1 or 100:1. In some preferred embodiments, the ratio of inserted first nucleic acid constructs to second nucleic acid constructs is from 1.2:1 to 1000:1, 500:1, 200:1 or 100:1. In some preferred embodiments, the ratio of inserted first nucleic acid constructs to second nucleic acid constructs is from 1.3:1 to 1000:1, 500:1, 200:1 or 100:1. In some preferred embodiments, the ratio of inserted first nucleic acid constructs to second nucleic acid constructs is from 1.4:1 to 1000:1, 500:1, 200:1 or 100:1. In some preferred embodiments, the ratio of inserted first nucleic acid constructs to second nucleic acid constructs is from 1.5:1 to 1000:1, 500:1, 200:1 or 100:1. In some preferred embodiments, the ratio of inserted first nucleic acid constructs to second nucleic acid constructs is from 2:1 to 1000:1, 500:1, 200:1 or 100:1. In some preferred embodiments, the ratio of inserted first nucleic acid constructs to second nucleic acid constructs is from 5:1 to 500:1. In some preferred embodiments, the ratio of inserted first nucleic acid constructs to second nucleic acid constructs is from 5:1 to 200:1. In some preferred embodiments, the ratio of inserted first nucleic acid constructs to second nucleic acid constructs is from 10:1 to 200:1. In some preferred embodiments, the ratio of first nucleic acid constructs to second nucleic acid constructs is from 10: 1 to 100: 1. In some preferred embodiments, the host cells comprise from 10 to 500 docking sites. In some preferred embodiments, the host cells comprise from 20 to 500 docking sites. In some preferred embodiments, the host cells comprise from 50 to 500 docking sites.In some preferred embodiments, one of the first and second proteins of interest is an enzyme. In some preferred embodiments, the first and second proteins of interest arc subunits of a multi-subunit protein. In some preferred embodiments, the first and second proteins are subunits of a viral particle. In some preferred embodiments, the nucleic acids of interest are viral nucleic acids, such as components of an adenoviral genome, AAV genome, or retroviral genome.In some preferred embodiments, only first nucleic acid constructs and second nucleic acid constructs are introduced into the host cell.In some preferred embodiments, the methods further comprise introducing a third nucleic acid construct encoding a third protein of interest or nucleic acid of interest into the host cell so that it is integrated at a ratio of first nucleic acid construct or second nucleic acid construct to third nucleic acid construct selected from the group consisting of at least 1.1:1, 1.2:1, 1.3:1, 1.4:1, 1.5:1 or 2:1, from 1.1 to 1000:1 or 100:1, from 1.2:1000:1 or 100:1, from 1.3 to 1000:1 or 100:1, from 1.4 to 1000:1 or 100:1, from 1.5 to 1000:1 or 100:1, from 2:1 to 1000:1 or 100:1, from 5:1 to 500:1 or 100:1, from 10:1 to 200:1 or 100:1, and from 10:1 to 100:1.In some preferred embodiments, the methods further comprise introducing a fourth nucleic acid construct encoding a fourth protein or nucleic acid of interest into the host cell so that it is integrated at a ratio of first nucleic acid construct, second nucleic acid construct, or third nucleic acid construct to the fourth nucleic construct selected from the group consisting of at least 1.1:1, 1.2:1, 1.3:1, 1.4:1, 1.5:1 or 2:1, from 1.1 to 1000:1 or 100:1, from 1.2:1000:1 or 100:1, from 1.3 to 1000:1 or 100:1, from 1.4 to 1000:1 or 100:1, from 1.5 to 1000:1 or 100:1, from 2:1 to 1000:1 or 100:1, from 5:1 to 500:1 or 100:1, from 10:1 to 200:1 or 100:1, and from 10:1 to 100:1.In some preferred embodiments, the methods further comprise introducing a fifth nucleic acid construct encoding a fifth protein or nucleic acid of interest into the host cell so that it is integrated at a ratio of first nucleic acid construct, second nucleic acid construct, third nucleic acid construct or fourth nucleic construct to the fifth nucleic acid construct selected from the group consisting of at least 1.1:1, 1.2:1, 1.3:1, 1.4:1, 1.5:1 or 2:1, from 1.1 to 1000:1 or 100:1, from 1.2:1000:1 or 100:1, from 1.3 to 1000:1 or 100:1, from 1.4 to 1000:1 or 100:1, from 1.5 to 1000:1 or 100:1, from 2:1 to 1000:1 or 100:1, from 5:1 to 500:1 or 100:1, from 10:1 to 200:1 or 100:1, and from 10:1 to 100:1.In some preferred embodiments, the at least first and second nucleic acid constructs further comprise at least the following elements in operable association in 5’ to 3’ order: an internal promoter sequence; a nucleic acid sequence encoding the first protein of interest or second protein that is operably linked to the internal promoter; and a poly A signal sequence.In some preferred embodiments, the at least first and second nucleic acid constructs comprise a selectable marker sequence. In some preferred embodiments, the at least first and second nucleic acid constructs comprise different selectable marker sequences. In some preferred embodiments, one of the first and second nucleic acid constructs comprises a selectable marker sequence and the other of the first and second nucleic acid constructs does not comprise a selectable marker sequence. In some preferred embodiments, the selectable marker sequences are 5’ to the internal promoter sequence and are operably linked to a 5’ promoter sequence.In some preferred embodiments, the nucleic acid construct comprises an extending packaging region (EPR) between the 5’ promoter and the selectable marker. In some preferred embodiments, the EPR comprises multiple potential Kozak sequences and / or ATG translation start sites.In some preferred embodiments, the promoter sequence is selected from the group consisting of SIN-LTR, SV40, EFla, E. coli lac, E. coli trp, phage lambda PL, phage lambda PR, T3, T7, cytomegalovirus (CMV) immediate early, herpes simplex virus (HSV) thymidine kinase, alpha-lactalbumin, and mouse metallothionein-I promoter sequences. In some preferred embodiments, the first promoter sequence is a weak promoter sequence. In some preferred embodiments, the first promoter sequence is not a retroviral LTR promoter. In some preferred embodiments, the integrated docking sites further comprise an exogenous promoter. In some preferred embodiments, the exogenous promoter is selected from the group consisting of SIN- LTR, SV40, EFla, E. coli lac, E. coli trp, phage lambda PL, phage lambda PR, T3, T7, cytomegalovirus (CMV) immediate early, herpes simplex vims (HSV) thymidine kinase, alphalactalbumin, and mouse metallothionein-I promoter sequences. In some preferred embodiments, the promoter is a retroviral LTR. In some preferred embodiments, the retroviral LTR is a SIN LTR.In some preferred embodiments, the nucleic acid expression constructs are provided in a vector. In some preferred embodiments, the vector is a plasmid vector. In some preferred embodiments, the vector is transiently introduced into the host cell.In some preferred embodiments, the host cell line comprises a nucleic acid construct encoding an enzyme that facilitates insertion of the nucleic acid expression construct at the dock site. In some preferred embodiments, the nucleic acid construct encoding an enzyme that facilitates insertion of the nucleic acid expression construct at the dock site is transiently introduced into the host cell. In some preferred embodiments, the nucleic acid construct encoding an enzyme that facilitates insertion of the nucleic acid expression construct at the dock site is provided in a vector. In some preferred embodiments, the vector is a plasmid vector. In some preferred embodiments, the ratio of the ratio of the nucleic acid constructs encoding an enzyme that facilitates insertion of the nucleic acid expression construct at the dock site to the nucleic acid expression constructs encoding a first protein of interest that are transiently introduced into the host cell line is from 1:1000 to 1:10. In some preferred embodiments, the enzyme is selected from the group consisting of an integrase, a recombinase, a nuclease and a nickase. In some preferred embodiments, the nucleic acid construct encoding an enzyme that facilitates insertion of the nucleic acid expression construct at the dock site is provided in a vector.In some preferred embodiments, the host cell genome comprises from 5 to 500 integrated docking sites, each docking site comprising at least one dock site insertion element. In some preferred embodiments, the host cell genome comprises from 5 to 250 integrated docking sites, each docking site comprising at least one dock site insertion element. In some preferred embodiments, the host cell genome comprises from 5 to 100 integrated docking sites, each docking site comprising at least one dock site insertion element. In some preferred embodiments, the integrated docking sites are independently positioned throughout the host cell genome.In some preferred embodiments, the dock site insertion element is targeted by enzyme selected from the group consisting of an integrase, a recombinase, a nuclease and a nickase. In some preferred embodiments, the dock site insertion element is selected from the group consisting of a recombinase dock site insertion element and a HDR dock site insertion element. In some preferred embodiments, the dock site insertion element is a recombinase dock site insertion element. In some preferred embodiments, the recombinase dock site insertion element comprises an attachment site (att). In some preferred embodiments, the attachment site (att) is selected from the group consisting of attB and attP and attR and attL. In some preferred embodiments, the recombinase dock site insertion element comprises a LoxP sequence. In somepreferred embodiments, the recombinase dock site insertion element is a Flp Recombination Target (FRT) site. In some preferred embodiments, is a HDR dock site insertion clement. In some preferred embodiments, the HDR dock site insertion element comprises one or two dock site homology arms. In some preferred embodiments, the HDR dock site insertion element further comprises one or more sequences homologous to a guide RNA sequence. In some preferred embodiments, the dock site homology arms are from about 30 to 1000 bases in length. In some preferred embodiments, the integrase dock site insertion element comprises an AAVS1 safe harbor locus sequence.In some preferred embodiments, each docking site is flanked by exogenous integrating vector sequences. In some preferred embodiments, the exogenous integrating vector sequences are selected from the group consisting of viral vector sequences and transposon vector sequences.In some preferred embodiments, the docking sites each further comprise a sequence encoding a selectable maker operably linked to a promoter.In some preferred embodiments, the dock site insertion element is positioned to facilitate cassette exchange. In some preferred embodiments, each docking site comprises two dock site insertion elements. In some preferred embodiments, the two dock site insertion elements are positioned to facilitate cassette exchange. In some preferred embodiments, the two dock site insertion elements flank sequences encoding a selectable marker, an enzyme, or a combination thereof.In some preferred embodiments, the nucleic acid expression constructs further comprise a signal peptide sequence operably linked to the first protein of interest. In some preferred embodiments, the signal peptide sequence is selected from the group consisting of tissue plasminogen activator, human growth hormone, lactoferrin, alpha-casein and alpha-lactalbumin signal peptide sequences. In some preferred embodiments, the nucleic acid expression constructs further comprise a protein purification marker sequence. In some preferred embodiments, the protein purification marker sequence is a hexahistidine tag or a hemagglutinin (HA) tag.In some preferred embodiments, the host cell is selected from the group consisting of Chinese Hamster Ovary (CHO) cells, HEK 293 cells, CAP cells, bovine mammary epithelial cells, monkey kidney CV1 line transformed by SV40, baby hamster kidney cells, mouse sertoli cells, monkey kidney cells, African green monkey kidney cells, human cervical carcinoma cells,canine kidney cells, buffalo rat liver cells, human lung cells, human liver cells, mouse mammary tumor, TRI cells, MRC 5 cells, FS4 cells, rat fibroblasts, MDBK cells, VERO cells, NSO cells, and human hepatoma line cells. In some preferred embodiments, the host cell is selected from the group consisting of Chinese Hamster Ovary (CHO) cells, HEK 293 cells and CAP cells. In some preferred embodiments, the host cell line is a GS knockout cell line. In some preferred embodiments, the host cell line is a DHFR knockout cell line.In some preferred embodiments, the present invention provides a cell culture comprising host cells made the methods described above.In some preferred embodiments, the present invention provides processes for producing a protein of interest comprising culturing host cells made by the methods described above under conditions that the protein of interest is expressed and purifying the protein of interest from the host cell culture. In some preferred embodiments, the host cells are grown in a medium comprising an inhibitor of the selectable marker. In some preferred embodiments, the selectable marker is GS and the inhibitor is phosphinothricin or methionine sulphoximine (Msx). In some preferred embodiments, the selectable marker is DHFR and the inhibitor is methotrexate.In some preferred embodiments, the present invention provides a host cell (or cells) comprising: a plurality of docking sites integrated into the genome of the host cell, each docking site comprising at least one dock site insertion element; at least integrated first nucleic acid constructs comprising at least one insertion element compatible with the dock site insertion element and encoding a first protein of interest, and at least integrated second nucleic acid constructs comprising at least one insertion element compatible with the dock site insertion and encoding a second protein of interest wherein the at least integrated first nucleic acid constructs and the at least second integrated nucleic acid constructs are integrated at the plurality of docking sites at a ratio of first nucleic acid constructs to second nucleic acid constructs of at least 1 : 1. In some preferred embodiments, the ratio of first nucleic acid constructs to second nucleic acid constructs is from 1.1:1 to 100:1 or 1000:1. In some preferred embodiments, the ratio of first nucleic acid constructs to second nucleic acid constructs is from 1.2:1 to 100:1 or 1000:1. In some preferred embodiments, the ratio of first nucleic acid constructs to second nucleic acid constructs is from 1.3:1 to 100:1 or 1000:1. In some preferred embodiments, the ratio of first nucleic acid constructs to second nucleic acid constructs is from 1.4:1 to 100:1 or 1000:1. In some preferred embodiments, the ratio of first nucleic acid constructs to second nucleic acidconstructs is from 1.5: 1 to 100: 1 or 1000:1 . In some preferred embodiments, the ratio of first nucleic acid constructs to second nucleic acid constructs is from 2:1 to 100:1 or 1000:1. In some preferred embodiments, the ratio of first nucleic acid constructs to second nucleic acid constructs is from 5:1 to 100:1 or 500:1. In some preferred embodiments, the ratio of first nucleic acid constructs to second nucleic acid constructs is from 10:1 to 100:1 or 200:1. In some preferred embodiments, the ratio of first nucleic acid constructs to second nucleic acid constructs is from 10:1 to 100:1.In some preferred embodiments, one of the first and second proteins of interest is a an enzyme. In some preferred embodiments, the first and second proteins of interest are subunits of a multi-subunit protein. In some preferred embodiments, the first and second proteins are subunits of a viral particle.In some preferred embodiments, the host cell further comprises a third integrated nucleic acid construct encoding a third protein of interest at a ratio of first nucleic acid construct or second nucleic acid construct to third nucleic acid construct selected from the group consisting of at least 1.1:1, 1.2:1, 1.3:1, 1.4:1, 1.5:1 or 2:1, from 1.1 to 1000:1 or 100:1, from 1.2:1000:1 or 100:1, from 1.3 to 1000:1 or 100:1, from 1.4 to 1000:1 or 100:1, from 1.5 to 1000:1 or 100:1, from 2:1 to 1000:1 or 100:1, from 5:1 to 500:1 or 100:1, from 10:1 to 200:1 or 100:1, and from 10:1 to 100:1. In some preferred embodiments, the host cell further comprises a fourth nucleic integrated acid construct encoding a fourth protein of interest at a ratio of first nucleic acid construct, second nucleic acid construct, or third nucleic acid construct to the fourth nucleic construct selected from the group consisting of at least 1.1:1, 1.2:1, 1.3:1, 1.4:1, 1.5:1 or 2:1, from 1.1 to 1000:1 or 100:1, from 1.2:1000:1 or 100:1, from 1.3 to 1000:1 or 100:1, from 1.4 to 1000:1 or 100:1, from 1.5 to 1000:1 or 100:1, from 2:1 to 1000:1 or 100:1, from 5:1 to 500:1 or 100:1, from 10:1 to 200:1 or 100:1, and from 10:1 to 100:1. In some preferred embodiments, the host cell further comprises a fifth integrated nucleic acid construct encoding a fifth protein of interest at a ratio of first nucleic acid construct, second nucleic acid construct, third nucleic acid construct or fourth nucleic construct to the fifth nucleic acid construct selected from the group consisting of at least 1.1:1, 1.2:1, 1.3:1, 1.4:1, 1.5:1 or 2:1, from 1.1 to 1000:1 or 100:1, from 1.2:1000:1 or 100:1, from 1.3 to 1000:1 or 100:1, from 1.4 to 1000:1 or 100:1, from 1.5 to 1000:1 or 100:1, from 2:1 to 1000:1 or 100:1, from 5:1 to 500:1 or 100:1, from 10:1 to 200:1 or 100:1, and from 10:1 to 100:1.In some preferred embodiments, the at least first and second nucleic acid constructs further comprise at least the following elements in operable association in 5’ to 3’ order: an internal promoter sequence; a nucleic acid sequence encoding the first protein of interest or second protein that is operably linked to the internal promoter; and a poly A signal sequence.In some preferred embodiments, the at least first and second nucleic acid constructs comprise a selectable marker sequence. In some preferred embodiments, the at least first and second nucleic acid constructs comprise different selectable marker sequences. In some preferred embodiments, one of the first and second nucleic acid constructs comprises a selectable marker sequence and the other of the first and second nucleic acid constructs does not comprise a selectable marker sequence. In some preferred embodiments, the selectable marker sequences are 5’ to the internal promoter sequence and are operably linked to a 5’ promoter sequence.In some preferred embodiments, the nucleic acid construct comprises an extending packaging region (EPR) between the 5’ promoter and the selectable marker. In some preferred embodiments, the EPR comprises multiple potential Kozak sequences and / or ATG translation start sites. In some preferred embodiments, the promoter sequence is selected from the group consisting of SIN-LTR, SV40, EFla, E. coli lac, E. coli trp, phage lambda PL, phage lambda PR, T3, T7, cytomegalovirus (CMV) immediate early, herpes simplex virus (HSV) thymidine kinase, alpha-lactalbumin, and mouse metallothionein-I promoter sequences. In some preferred embodiments, the first promoter sequence is a weak promoter sequence. In some preferred embodiments, the first promoter sequence is not a retroviral LTR promoter. In some preferred embodiments, the integrated docking sites further comprise an exogenous promoter. In some preferred embodiments, the exogenous promoter is selected from the group consisting of SIN- LTR, SV40, EFla, E. coli lac, E. coli trp, phage lambda PL, phage lambda PR, T3, T7, cytomegalovirus (CMV) immediate early, herpes simplex virus (HSV) thymidine kinase, alphalactalbumin, and mouse metallothionein-I promoter sequences. In some preferred embodiments, the promoter is a retroviral LTR. In some preferred embodiments, the retroviral LTR is a SIN LTR.In some preferred embodiments, the nucleic acid expression constructs are provided in a vector. In some preferred embodiments, the vector is a plasmid vector.In some preferred embodiments, the host cell further comprises a nucleic acid construct encoding an enzyme that facilitates insertion of the nucleic acid expression construct at the docksite. Tn some preferred embodiments, the nucleic acid construct encoding an enzyme that facilitates insertion of the nucleic acid expression construct at the dock site is provided in a vector. In some preferred embodiments, the vector is a plasmid vector. In some preferred embodiments, the ratio of the nucleic acid constructs encoding an enzyme that facilitates insertion of the nucleic acid expression construct at the dock site to the nucleic acid expression constructs encoding a first protein of interest that are transiently introduced into the host cell line is from 1:1000 to 1:10. In some preferred embodiments, the enzyme is selected from the group consisting of an integrase, a recombinase, a nuclease and a nickase. In some preferred embodiments, the nucleic acid construct encoding an enzyme that facilitates insertion of the nucleic acid expression construct at the dock site is provided in a vector.In some preferred embodiments, the host cell genome comprises from 5 to 500 integrated docking sites, each docking site comprising at least one dock site insertion element. In some preferred embodiments, the host cell genome comprises from 5 to 250 integrated docking sites, each docking site comprising at least one dock site insertion element. In some preferred embodiments, the host cell genome comprises from 5 to 100 integrated docking sites, each docking site comprising at least one dock site insertion element. In some preferred embodiments, wherein the integrated docking sites are independently positioned throughout the host cell genome.In some preferred embodiments, the dock site insertion element is targeted by enzyme selected from the group consisting of an integrase, a recombinase, a nuclease and a nickase. In some preferred embodiments, the dock site insertion element is selected from the group consisting of a recombinase dock site insertion element and a HDR dock site insertion element. In some preferred embodiments, the dock site insertion element is a recombinase dock site insertion element. In some preferred embodiments, the recombinase dock site insertion element comprises an attachment site (att). In some preferred embodiments, the attachment site (att) is selected from the group consisting of attB and attP and attR and attL. In some preferred embodiments, the recombinase dock site insertion element comprises a LoxP sequence. In some preferred embodiments, the recombinase dock site insertion element is a Flp Recombination Target (FRT) site. In some preferred embodiments, the dock site insertion element is a HDR dock site insertion element. In some preferred embodiments, the HDR dock site insertion element comprises one or two dock site homology arms. In some preferred embodiments, theHDR dock site insertion element further comprises one or more sequences homologous to a guide RNA sequence. In some preferred embodiments, the dock site homology arms arc from about 30 to 1000 bases in length. In some preferred embodiments, the integrase dock site insertion element comprises an AAVS1 safe harbor locus sequence.In some preferred embodiments, each docking site is flanked by exogenous integrating vector sequences. In some preferred embodiments, the exogenous integrating vector sequences are selected from the group consisting of viral vector sequences and transposon vector sequences.In some preferred embodiments, the docking sites each further comprise a sequence encoding a selectable maker operably linked to a promoter.In some preferred embodiments, the host cell further comprises an expression construct encoding an exogenous enzyme selected from the group consisting of an integrase, a recombinase, a nuclease and a nickase. In some preferred embodiments, the expression construct encoding an exogenous enzyme selected from the group consisting of an integrase, a recombinase, a nuclease and a nickase is provided in an episomal expression vector. In some preferred embodiments, the expression construct encoding an exogenous enzyme selected from the group consisting of an integrase, a recombinase, a nuclease and a nickase is integrated into the host cell genome.In some preferred embodiments, the dock site insertion element is positioned to facilitate cassette exchange. In some preferred embodiments, the docking site comprises two dock site insertion elements. In some preferred embodiments, the two dock site insertion elements are positioned to facilitate cassette exchange. In some pre I erred embodiments, the two dock site insertion elements flank sequences encoding a selectable marker, an enzyme, or a combination thereof.In some preferred embodiments, the nucleic acid expression constructs further comprise a signal peptide sequence operably linked to the first protein of interest. In some preferred embodiments, signal peptide sequence is selected from the group consisting of tissue plasminogen activator, human growth hormone, lactoferrin, alpha-casein and alpha-lactalbumin signal peptide sequences.In some preferred embodiments, the nucleic acid expression constructs further comprise a protein purification marker sequence. In some preferred embodiments, the protein purification marker sequence is a hexahistidine tag or a hemagglutinin (HA) tag.In some preferred embodiments, the host cell is selected from the group consisting of Chinese Hamster Ovary (CHO) cells, HEK 293 cells, CAP cells, bovine mammary epithelial cells, monkey kidney CV1 line transformed by SV40, baby hamster kidney cells, mouse sertoli cells, monkey kidney cells, African green monkey kidney cells, human cervical carcinoma cells, canine kidney cells, buffalo rat liver cells, human lung cells, human liver cells, mouse mammary tumor, TRI cells, MRC 5 cells, FS4 cells, rat fibroblasts, MDBK cells, VERO cells, NSO cells, and human hepatoma line cells. In some preferred embodiments, the host cell is selected from the group consisting of Chinese Hamster Ovary (CHO) cells, HEK 293 cells and CAP cells. In some preferred embodiments, the host cell line is a GS knockout cell line. In some preferred embodiments, the host cell line is a DHFR knockout cell line.In some preferred embodiments, the present invention provides a cell culture comprising the host cell(s) as described above.In some preferred embodiments, the present invention provides a process for producing a protein of interest comprising culturing host cells as described above under conditions that the protein(s) of interest are expressed and purifying the protein(s) of interest from the host cell culture. In some preferred embodiments, the host cells are grown in a medium comprising an inhibitor of the selectable marker. In some preferred embodiments, the selectable marker is GS and the inhibitor is phosphinothricin or methionine sulphoximine (Msx). In some preferred embodiments, the selectable marker is DHFR and the inhibitor is methotrexate.DESCRIPTION OF THE FIGURESFIG. 1A and B. Human DNase 1 nucleic acid sequence and the flanking DNA cloning junctions (SEQ ID NO:1) in the final retrovector expression construct (FIG 1A) and DNase amino acid sequence (SEQ ID NO:2, FIG. IB).FIG. 2. Map of starting Retrovector pCS-newMCS-WPRE (new ori), 6187 bp.FIG. 3. Map of Human DNase 1 “Pathway” Gene in GPEx® Vector pCS-CFSDl-WPRE (new ori), 6977 bp.FIG. 4. Map of plasmid 207attB-GS-h3E10LC-WPRE.FIG. 5. Map of plasmid 207 attB-GS-h3E10HC-WPREFIG. 6A-B. Sequence data for Pathway CDS in 215-pucl9attB287-GS-Pathway-WPRE- TKpa. FIG. 6A provides the nucleic acid sequence (SEQ ID NO:3) while FIG. 6B provides the amino acid sequence (SEQ ID NO:4).FIG. 7. Map of plasmid 215-pucl9attB287-GS-Pathway-WPRE-TKpaFIG. 8. SDS-PAGE results for pooled cells.FIG. 9. Graph showing viable cell density.FIG. 10. Graph showing percent viability of cells.FIG. 11. Graph showing antibody titer.FIG. 12. Graph showing protein concentration (mg / ml) vs. clone number.FIG. 13. Graph showing Ambrl5™ IVCD vs clone number.FIG. 14. Graph showing Ambrl5™ rQp vs clone number.FIG. 15 provides a sequence summary for Yourway Light Chain (LC) CDS in vectors GDD1008.0211 and GDD1008.0223.FIG. 16 provides a map of plasmid 223attB-GS-Yourway LC-WPRE, GDD1008.0223.FIG. 17 provides a sequence summary for Yourway Heavy Chain (HC) CDS in vectors GDD1008.0211 and GDD1008.0224.FIG. 18 provides a map of plasmid 224attB-GS-Yourway HC-WPRE, GDD1008.0224.FIG. 19 provides a map of plasmid 21 lattB287-GS-sC-YourwayHC-WPRE-hC-Int- YourwayLC, GDD1008.0211.FIG. 20 provides a graph showing cell viability curves after glutamine selection.FIG. 21 provides a graph showing pooled cell productivity of the different heavy and light chain ratios as well as the double chain construct (CT36-2).FIG. 22 provides a non-reduced and reduced SDS-PAGE of samples all of the heavy chain / light chain ratio cell pools as well as the single gene construct (Historic Yourway HWIL).DEFINITIONSTo facilitate understanding of the invention, a number of terms are defined below.As used herein, the term "host cell" refers to any eukaryotic cell (e.g., mammalian cells, avian cells, amphibian cells, plant cells, fish cells, and insect cells), whether located in vitro or in vivo.As used herein, the term "cell culture" refers to any in vitro culture of cells. Included within this term are continuous cell lines (e.g., with an immortal phenotype), primary cell cultures, finite cell lines (e.g., non-transformed cells), and any other cell population maintained in vitro, including oocytes and embryos.As used herein, the term "vector" refers to any genetic element, such as a plasmid, phage, transposon, cosmid, chromosome, virus, virion, etc., which is capable of replication when associated with the proper control elements and which can transfer gene sequences between cells. Thus, the term includes cloning and expression vehicles, as well as viral vectors.As used herein, the term “genome” refers to the genetic material (e.g., chromosomes) of an organism.The term "nucleotide sequence of interest" refers to any nucleotide sequence (e.g., RNA or DNA), the manipulation of which may be deemed desirable for any reason (e.g., treat disease, confer improved qualities, expression of a protein of interest in a host cell, expression of a ribozyme, etc.), by one of ordinary skill in the art. Such nucleotide sequences include, but are not limited to, coding sequences of structural genes (e.g., reporter genes, selection marker genes, oncogenes, drug resistance genes, growth factors, etc.), and non-coding regulatory sequences which do not encode an mRNA or protein product (e.g., promoter sequence, polyadenylation sequence, termination sequence, enhancer sequence, etc.).As used herein, the term “product of interest” refers to a protein or nucleic acid product, such as a viral backbone or genome, encoded by a nucleic acid of interest. In some preferred embodiments, a plurality of products of interest are expressed in a host cell.As used herein, the term “protein of interest” refers to a protein encoded by a nucleic acid of interest.As used herein, the terms "nucleic acid molecule encoding," "DNA sequence encoding," "DNA encoding," "RNA sequence encoding," and "RNA encoding" refer to the order or sequence of deoxyribonucleotides or ribonucleotides along a strand of deoxyribonucleic acid or ribonucleic acid. The order of these deoxyribonucleotides or ribonucleotides determines theorder of amino acids along the polypeptide (protein) chain. The DNA or RNA sequence thus codes for the amino acid sequence.The term "promoter," "promoter element," or "promoter sequence" as used herein, refers to a DNA sequence which when ligated to a nucleotide sequence of interest is capable of controlling the transcription of the nucleotide sequence of interest into mRNA. A promoter is typically, though not necessarily, located 5' (z.e., upstream) of a nucleotide sequence of interest whose transcription into mRNA it controls, and provides a site for specific binding by RNA polymerase and other transcription factors for initiation of transcription.Transcriptional control signals in eukaryotes comprise "promoter" and "enhancer" elements. Promoters and enhancers consist of short arrays of DNA sequences that interact specifically with cellular proteins involved in transcription (Maniatis et al., Science 236:1237

[1987] ). Promoter and enhancer elements have been isolated from a variety of eukaryotic sources including genes in yeast, insect and mammalian cells, and viruses (analogous control elements, i.e., promoters, are also found in prokaryotes). The selection of a particular promoter and enhancer depends on what cell type is to be used to express the protein of interest. Some eukaryotic promoters and enhancers have a broad host range while others are functional in a limited subset of cell types (for review see, Voss et al., Trends Biochem. Sci., 11:287

[1986] ; and Maniatis et al., supra). For example, the SV40 early gene enhancer is very active in a wide variety of cell types from many mammalian species and has been widely used for the expression of proteins in mammalian cells (Dijkema et al., EMBO J. 4:761

[1985] ). Two other examples of promoter / enhancer elements active in a broad range of mammalian cell types are those from the human elongation factor la gene (Uetsuki et al., J. Biol. Chem., 264:5791

[1989] ; Kim et al., Gene 91:217

[1990] ; and Mizushima and Nagata, Nuc. Acids. Res., 18:5322

[1990] ) and the long terminal repeats of the Rous sarcoma virus (Gorman et al., Proc. Natl. Acad. Sci. USA 79:6777

[1982] ) and the human cytomegalovirus (Boshart et al., Cell 41:521

[1985] ).As used herein, the term "promoter / enhancer" denotes a segment of DNA which contains sequences capable of providing both promoter and enhancer functions (i.e., the functions provided by a promoter element and an enhancer element, see above for a discussion of these functions). For example, the long terminal repeats of retroviruses contain both promoter and enhancer functions. The enhancer / promoter may be "endogenous" or "exogenous" or "heterologous." An "endogenous" enhancer / promoter is one that is naturally linked with a givengene in the genome. An "exogenous" or "heterologous" enhancer / promoter is one that is placed in juxtaposition to a gene by means of genetic manipulation (i.e., molecular biological techniques such as cloning and recombination) such that transcription of that gene is directed by the linked enhancer / promoter.As used herein, the term “long terminal repeat” of "LTR" refers to transcriptional control elements located in or isolated from the U3 region 5' and 3' of a retroviral genome. As is known in the art, long terminal repeats may be used as control elements in retroviral vectors, or isolated from the retroviral genome and used to control expression from other types of vectors.As used herein, the terms "complementary" or "complementarity" are used in reference to polynucleotides (i.e., a sequence of nucleotides) related by the base-pairing rules. For example, the sequence ”5'-A-G-T-3'," is complementary to the sequence ”3'-T-C-A-5'." Complementarity may be "partial," in which only some of the nucleic acids' bases are matched according to the base pairing rules. Or, there may be "complete" or "total" complementarity between the nucleic acids. The degree of complementarity between nucleic acid strands has significant effects on the efficiency and strength of hybridization between nucleic acid strands. This is of particular importance in amplification reactions, as well as detection methods that depend upon binding between nucleic acids.The terms "homology" and "percent identity" when used in relation to nucleic acids refers to a degree of complementarity. There may be partial homology (i.e., partial identity) or complete homology (i.e., complete identity). A partially complementary sequence is one that at least partially inhibits a completely complementary sequence from hybridizing to a target nucleic acid sequence and is re I erred to using the functional term "substantially homologous." The inhibition of hybridization of the completely complementary sequence to the target sequence may be examined using a hybridization assay (Southern or Northern blot, solution hybridization and the like) under conditions of low stringency. A substantially homologous sequence or probe (i.e., an oligonucleotide which is capable of hybridizing to another oligonucleotide of interest) will compete for and inhibit the binding (i.e., the hybridization) of a completely homologous sequence to a target sequence under conditions of low stringency. This is not to say that conditions of low stringency are such that non-specific binding is permitted; low stringency conditions require that the binding of two sequences to one another be a specific (i.e., selective) interaction. The absence of non-specific binding may be tested by the use of a second targetwhich lacks even a partial degree of complementarity (e.g., less than about 30% identity); in the absence of non-specific binding the probe will not hybridize to the second non-complcmcntary target.The terms "in operable combination," "in operable order," and "operably linked" as used herein refer to the linkage of nucleic acid sequences in such a manner that a nucleic acid molecule capable of directing the transcription of a given gene and / or the synthesis of a desired protein molecule is produced. The term also refers to the linkage of amino acid sequences in such a manner so that a functional protein is produced.As used herein, the term “selectable marker” refers to a gene that encodes an enzymatic activity or other protein that confers the ability to grow in medium lacking what would otherwise be an essential nutrient; in addition, a selectable marker may confer resistance to an antibiotic or drug upon the cell in which the selectable marker is expressed.As used herein, the term “retrovirus" refers to a retroviral particle which is capable of entering a cell (z.e., the particle contains a membrane-associated protein such as an envelope protein or a viral G glycoprotein which can bind to the host cell surface and facilitate entry of the viral particle into the cytoplasm of the host cell) and integrating the retroviral genome (as a double-stranded provirus) into the genome of the host cell. The term "retrovirus" encompasses Oncovirinae (e.g., Moloney murine leukemia virus (MoMLV), Moloney murine sarcoma virus (MoMSV), and Mouse mammary tumor virus (MMTV), Spumavirinae, and Lentiviridae (e.g., Human immunodeficiency virus, Simian immunodeficiency virus, Equine infection anemia virus, and Caprine arthritis-encephalitis virus; See, e.g., U.S. Pat. Nos. 5,994,136 and 6,013,516, both of which are incorporated herein by reference).As used herein, the term "retroviral vector" refers to a retrovirus that has been modified to express a gene of interest. Retroviral vectors can be used to transfer genes efficiently into host cells by exploiting the viral infectious process. Foreign or heterologous genes cloned (i.e., inserted using molecular biological techniques) into the retroviral genome can be delivered efficiently to host cells that are susceptible to infection by the retrovirus. Through well-known genetic manipulations, the replicative capacity of the retroviral genome can be destroyed. The resulting replication-defective vectors can be used to introduce new genetic material to a cell but they are unable to replicate. A helper virus or packaging cell line can be used to permit vector particle assembly and egress from the cell. Such retroviral vectors comprise a replication-deficient retroviral genome containing a nucleic acid sequence encoding at least one gene of interest (z.e., a polycistronic nucleic acid sequence can encode more than one gene of interest), a 5' retroviral long terminal repeat (5' LTR); and a 3' retroviral long terminal repeat (3' LTR).As used herein, the term “lentivirus vector” refers to retroviral vectors derived from the Lentiviridae family (e.g., human immunodeficiency virus, simian immunodeficiency virus, equine infectious anemia virus, and caprine arthritis-encephalitis virus) that are capable of integrating into non-dividing cells (See, e.g., U.S. Pat. Nos. 5,994,136 and 6,013,516, both of which are incorporated herein by reference).As used herein, the term “transposon” refers to transposable elements (e.g., Tn5, Tn7, and TnlO) that can move or transpose from one position to another in a genome. In general, the transposition is controlled by a transposase. The term "transposon vector," as used herein, refers to a vector encoding a nucleic acid of interest flanked by the terminal ends of transposon. Examples of transposon vectors include, but are not limited to, those described in U.S. Pat. Nos. 6,027,722; 5,958,775; 5,968,785; 5,965,443; and 5,719,055, all of which are incorporated herein by reference.As used herein, the term “adeno-associated virus (AAV) vector” refers to a vector derived from an adeno-associated virus serotype, including without limitation, AAV-1, AAV-2, AAV-3, AAV-4, AAV-5, AAVX7, etc. AAV vectors can have one or more of the AAV wildtype genes deleted in whole or pail, preferably the rep and / or cap genes, but retain functional flanking ITR sequences.AAV vectors can be constructed using recombinant techniques that are known in the art to include one or more heterologous nucleotide sequences flanked on both ends (5' and 3') with functional AAV ITRs. In the practice of the invention, an AAV vector can include at least one AAV ITR and a suitable promoter sequence positioned upstream of the heterologous nucleotide sequence and at least one AAV ITR positioned downstream of the heterologous sequence. A "recombinant AAV vector plasmid" refers to one type of recombinant AAV vector wherein the vector comprises a plasmid. As with AAV vectors in general, 5' and 3' ITRs flank the selected heterologous nucleotide sequence.As used herein, the term “adenoviral vector” refers to a non-enveloped double-stranded DNA vector comprising an adenovirus backbone.As used herein, the term "purified" refers to molecules, either nucleic or amino acid sequences, that arc removed from their normal environment, isolated or separated. An "isolated nucleic acid sequence" is therefore a purified nucleic acid sequence. "Substantially purified" molecules are at least 60% free, preferably at least 75% free, and more preferably at least 90% free from other components with which they are normally associated.ABBREVIATIONSAmpR= bacterial ampicillin resistance gene attB= Bacterial Attachment Site attP= Phage Attachment Site attR= Recombined Upstream Attachment SiteBackbone=Plasmid BackboneCDS= Coding SequenceEPR=MMLV Extended Packaging RegionGO = Gene Copy IndexGS= Glutamine SynthetaseH or HC= Heavy Chain hCMV= Human Cytomegalovirus immediate-early Promoter1= intronL or LC= Light ChainMoMuSV 5’LTR= Moloney Murine Sarcoma Virus 5’ Long Terminal RepeatNeo= Neomycin resistance genePA or PolyA= Polyadenylation signalProV SIN-LTR= Proviral Self- Inactivating Long Terminal Repeat sCMV= Simian Cytomegalovirus immediate-early PromoterSDS-PAGE= Sodium Dodecyl Sulphate- Polyacrylamide Gel ElectrophoresisSIN-3’LTR= Self-Inactivation 3’ Long Terminal RepeatSV40= Simian Virus 40TK= Thymidine KinaseUTR= Untranslated RegionW or WPRE= Woodchuck Post-transcriptional Regulatory ElementDETAILED DESCRIPTION OF THE INVENTIONIn some embodiments, the present invention provides host cells that allow for coexpression DNase with a product of interest to improve expression and / or purification of the product of interest from the host cell culture. In other embodiments, the present invention provides host cells and associated methods that allow for expression of two or more products of interest at varying ratios in a host cell.In some particularly preferred embodiments, an expression construct encoding DNase and an expression construct(s) encoding one or more additional gene(s) of interest are introduced into host cells at defined ratios. In some particularly preferred embodiments, the host cell lines contain multiple dock sites for insertion of the nucleic acid constructs. Cell lines containing multiple dock sites and expression constructs for use with the cells are described in PCT / US21 / 35403 and PCT / US21 / 35404, both of which are incorporated by reference herein in their entirety.The present invention solves multiple problems by providing host cells that allow for coexpression of DNase with one or more proteins of interest. In one aspect, co-expression of DNase with one or more products of interest allows for more efficient removal of host cell DNA from pharmaceutical preparations. In another aspect, co-expression of DNase with one or more products of interest allows for more efficient production of the product of interest, especially where the product of interest may interact with host cell DNA. In both aspects, the inventors have surprisingly found that DNase can be stably co-expressed in host cells lines. The inventors have also found that when two proteins are expressed in the system, the yield of one or both of the proteins (or for example, a multi-subunit protein comprising the two proteins such as an antibody) can be improved by varying the ratio of the expression constructs.Accordingly, in some embodiments, the present invention provides a eukaryotic host cell comprising a first exogenous nucleic acid sequence encoding DNase 1 operably linked to a promoter sequence and at least a second exogenous nucleic acid sequence encoding a first product of interest operably linked to a promoter sequence, wherein the first sequence encoding DNasel and the at least second exogenous encoding the first product of interest are co-expressed in the host cell. In some preferred embodiments, the present invention provides a host cell culture comprising the host cells.In some preferred embodiments, the host cells comprise multiple exogenous nucleic acids encoding multiple products of interest, for example, 2, 3, 4, 5, 6, 7, 8, 9 or 10 products of interest.The present invention is not limited to the use of any particular DNase. In some preferred embodiments, the DNase sequence incorporated into the construct encodes a DNasel i.e., a protein having DNase activity) having at least 80% sequence identity to SEQ ID NO:2. In some preferred embodiments, the DNase sequence incorporated into the construct encodes a DNase having at least 90% sequence identity to SEQ ID NO:2. In some preferred embodiments, the DNase sequence incorporated into the construct encodes a DNase having at least 95% sequence identity to SEQ ID NO:2. In some preferred embodiments, the DNase sequence incorporated into the construct encodes a DNase having at least 98% sequence identity to SEQ ID NO:2. In some preferred embodiments, the DNase sequence incorporated into the construct encodes a DNase having SEQ ID NO:2. In other preferred embodiments, the DNase sequence incorporated into the construct has at least 80% sequence identity with the DNase encoding portions of SEQ ID NO: 1. In some preferred embodiments, the DNase sequence incorporated into the construct has at least 90% sequence identity with the DNase encoding portions of SEQ ID NO:1. In some preferred embodiments, the DNase sequence incorporated into the construct has at least 95% sequence identity with the DNase encoding portions of SEQ ID NO:1. In some preferred embodiments, the DNase sequence incorporated into the construct has at least 98% sequence identity with the DNase encoding portions of SEQ ID NO:1. In some preferred embodiments, the DNase sequence incorporated into the construct is SEQ ID NO:1.The present invention is not limited to the expression of any particular product of interest. In some preferred embodiments, the product or products of interest are proteins. In some preferred embodiments, one protein of interest may be co-expressed with DNase. In some preferred embodiments, two or more proteins of interest may be co-expressed with DNase. In some preferred embodiments, two proteins of interest may be co-expressed with DNase. In some preferred embodiments, three proteins of interest may be co-expressed with DNase. In some preferred embodiments, four proteins of interest may be co-expressed with DNase. In some preferred embodiments, five proteins of interest may be co-expressed with DNase. In some preferred embodiments, the protein of interest is selected from the group consisting of an Fc- fusion protein, an enzyme, an albumin fusion, a growth factor, a protein receptor, a single chainantibody (scFv), a single chain-Fc (scFv-Fc), a diabody, and minibody (scFv-CH3), Fab, single chain Fab (scFab), an immunoglobulin heavy chain, and an immunoglobulin light chain and other antigen binding proteins. In general, the protein or proteins of interest may be any pharmaceutical or industrial protein for which expression and production via a host culture is desired. In some preferred embodiments, the protein of interest is a biopharmaceutical protein. In some preferred embodiments, the protein of interest is an immunoglobulin heavy and / or light chain. In some embodiments, the protein of interest is an immunoglobulin fragment or single chain antibody. In some preferred embodiments, where expression of an immunoglobulin is desired, the host cell further comprises a third exogenous nucleic acid sequence encoding a second protein of interest operably linked to a promoter sequence and a secretion signal sequence, wherein the first protein of interest is an immunoglobulin light chain sequence and the second protein of interest is an immunoglobulin heavy chain sequence.In some preferred embodiments, the product or products of interest are nucleic acid product, for example nucleic acids that are or form part of a virus or exosome. In some preferred embodiments, the viral nucleic acids are selected from the group consisting of retroviral nucleic acids, lentiviral nucleic acids, adenoviral nucleic acids, and adeno-associated virus (AAV) nucleic acids.. The viral nucleic acids may preferably encode a viral backbone, genome or other nucleic acid needed for viral replication or packaging. In some preferred embodiments, product of interest is a retroviral backbone or lentiviral backbone. In some preferred embodiments, the nucleic acid of interest may be co-expressed with a protein of interest, for example a viral capsid or envelope protein, such as a retrovirus, lentivirus or AAV capsid or envelope protein. In some preferred embodiments, where the viral backbone is a lentiviral or retroviral backbone, the capsid or envelope protein may be a protein that allows pseudotyping such as the VSV-G protein.In some preferred embodiments, the product of interest is one or more AAV nucleic acids. In some preferred embodiments, the AAV nucleic acids may preferably include a pHelper sequence and / or an adenoviral backbone sequence with or without a gene of interest. In some preferred embodiments, the AAV nucleic acid sequence(s) may be co-expressed with sequences of interest encoding AAV Rep and Cap proteins, which can, for example, be the first and second or second and third proteins or products of interest in the embodiments described above or below. Suitable AAV sequences are described, for example, in U.S. Pat. Nos. 5,622,856;5,945,335; 6,001 ,650; 6,004,797; 6,027,931 ; 6,376,237; 6,365,403; 6,482,633; 6,897,063; 7,037,713; 7,638,120; 6,759,237; 8,906,675; 7,282,199; 7,906,111; and 9,790,472, each of which incorporated herein by reference in its entirety.In some preferred embodiments, one nucleic acid product of interest may be co-expressed with DNase. In some preferred embodiments, two or more nucleic acid products of interest may be co-expressed with DNase. In some preferred embodiments, two nucleic acid products of interest may be co-expressed with DNase. In some preferred embodiments, three nucleic acid products of interest may be co-expressed with DNase. In some preferred embodiments, four nucleic acid products of interest may be co-expressed with DNase. In some preferred embodiments, five nucleic acid products of interest may be co-expressed with DNase. In some preferred embodiments, the first exogenous nucleic acid sequence encoding a DNase and the at least a second exogenous nucleic acid sequence encoding a product of interest are stably integrated into the genome of the host cell. In some preferred embodiments, the first exogenous nucleic acid sequence and at least a second exogenous nucleic acid sequence are stably integrated into the genome of the host cell at a docking site as described in further detail below.The host cells may preferably comprise a plurality of integrated sequences. In some preferred embodiments, from 10 to 500 copies each of the first exogenous nucleic acid sequence and at least a second exogenous nucleic acid sequence are stably integrated into the genome of the host cell. In some preferred embodiments, from 10 to 200 copies each of the first exogenous nucleic acid sequence and at least a second exogenous nucleic acid sequence are stably integrated into the genome of the host cell.In other preferred embodiments, the present invention provides methods for integrating two or more (e.g., 2, 3, 4, 5, 6, 7, 8, 9, or 10 or more) nucleic acid constructs, each preferably encoding a product or products of interest, into a host cell line at a desired ratios, and host cells produced by such processes. These embodiments are not limited to expression of DNase (but may include expression of DNase) and extend to the expression of any two or more products of interest, such as proteins or nucleic acids, at desired ratios.The ratio of the integrated sequences in the host cell may be varied. In some preferred embodiments, the ratio of the first exogenous nucleic acid sequence (e.g., DNase or an antibody heavy or light chain) to the second exogenous nucleic acid sequence (e.g., an antibody heavy or light chain or scFv) is from 1:1 to 1:10,000. In some preferred embodiments, the ratio of the firstexogenous nucleic acid sequence to the second exogenous nucleic acid sequence is from 1 :1 to 1:1000. In some preferred embodiments, the ratio of the first exogenous nucleic acid sequence to the second exogenous nucleic acid sequence is from 1:1 to 1:100. In some preferred embodiments, the ratio of the first exogenous nucleic acid sequence (e.g., DNase) to the second exogenous nucleic acid sequence is from 1:2 to 1:10,000. In some preferred embodiments, the ratio of the first exogenous nucleic acid sequence to the second exogenous nucleic acid sequence is from 1:2 to 1:1000. In some preferred embodiments, the ratio of the first exogenous nucleic acid sequence to the second exogenous nucleic acid sequence is from 1:2 to 1:200. In some preferred embodiments, the ratio of the first exogenous nucleic acid sequence to the second exogenous nucleic acid sequence is from 1:5 to 1:200. In some preferred embodiments, the ratio of the first exogenous nucleic acid sequence to the second exogenous nucleic acid sequence is from 1:10 to 1:200. In some preferred embodiments, the ratio of the first exogenous nucleic acid sequence to the second exogenous nucleic acid sequence is from 1:2 to 1:100. In some preferred embodiments, the ratio of the first exogenous nucleic acid sequence (e.g., DNase) to the second exogenous nucleic acid sequence is from 1:5 to 1:10,000. In some preferred embodiments, the ratio of the first exogenous nucleic acid sequence to the second exogenous nucleic acid sequence is from 1:5 to 1:1000. In some preferred embodiments, the ratio of the first exogenous nucleic acid sequence to the second exogenous nucleic acid sequence is from 1:5 to 1:100. In some preferred embodiments, the ratio of the first exogenous nucleic acid sequence to the second exogenous nucleic acid sequence is from 1:1 to 1:50. In some preferred embodiments, the ratio of the first exogenous nucleic acid sequence to the second exogenous nucleic acid sequence is from 1:2 to 1:50. In some preferred embodiments, the ratio of the first exogenous nucleic acid sequence to the second exogenous nucleic acid sequence is from 1:5 to 1:50. In some preferred embodiments, the ratio of the first exogenous nucleic acid sequence to the second exogenous nucleic acid sequence is from 1:1 to 1:20. In some preferred embodiments, the ratio of the first exogenous nucleic acid sequence to the second exogenous nucleic acid sequence is from 1:2 to 1:20. In some preferred embodiments, the ratio of the first exogenous nucleic acid sequence to the second exogenous nucleic acid sequence is from 1:5 to 1:20.As mentioned above, in some preferred embodiments, a second exogenous sequence (e.g., encoding an antibody heavy chain) and a third exogenous sequence (e.g., encoding an antibody light chain) are co-expressed with the first exogenous nucleic acid sequence (e.g.,DNase). In these embodiments, the ratios of the three integrated sequences in the host cell may also be varied. In some preferred embodiments, the ratio of the first exogenous sequence : second exogenous sequence : third exogenous sequence is from 1:1:1 to 1:100:100. In some preferred embodiments, the ratio of the first exogenous sequence : second exogenous sequence : third exogenous sequence is from 1:2:2 to 1:100:100. In some preferred embodiments, the ratio of the first exogenous sequence : second exogenous sequence : third exogenous sequence is from 1:5:5 to 1:100:100. In some preferred embodiments, the ratio of the first exogenous sequence : second exogenous sequence : third exogenous sequence is from 1:10:10 to 1:100:100. In some preferred embodiments, the ratio of the first exogenous sequence : second exogenous sequence : third exogenous sequence is from 1:1:1 to 1:50:50. In some preferred embodiments, the ratio of the first exogenous sequence : second exogenous sequence : third exogenous sequence is from 1:2:2 to 1:50:50. In some preferred embodiments, the ratio of the first exogenous sequence : second exogenous sequence : third exogenous sequence is from 1:5:5 to 1:50:500. In some preferred embodiments, the ratio of the first exogenous sequence : second exogenous sequence : third exogenous sequence is from 1:10:10 to 1:50:50. In some preferred embodiments, the ratio of the first exogenous sequence : second exogenous sequence : third exogenous sequence is from 1:1:1 to 1:20:20. In some preferred embodiments, the ratio of the first exogenous sequence : second exogenous sequence : third exogenous sequence is from 1:2:2 to 1:20:20. In some preferred embodiments, the ratio of the first exogenous sequence : second exogenous sequence : third exogenous sequence is from 1:5:5 to 1:20:20. In some preferred embodiments, the ratio of the first exogenous sequence : second exogenous sequence : third exogenous sequence is from 1:10:10 to 1:20:20. In some preferred embodiments, the ratio of the first exogenous sequence : second exogenous sequence : third exogenous sequence is from 1:2:2 to 1:15:15. In some preferred embodiments, the ratio of the first exogenous sequence : second exogenous sequence : third exogenous sequence is from 1:5:5 to 1:15:15.Similar ratios may be employed when 4, 5, 6, 7, 8, 9 or 10 products of interest are expressed.In further aspects, the present invention may be utilized to express any two or more proteins or nucleic acids from corresponding nucleic acid constructs that can be incorporated into the docking sites in the host cell. In some embodiments, at least one of the proteins is an enzyme. In other preferred embodiments, the first and second proteins of interest are subunits of a multi-subunit protein, such as the heavy and light chains of an immunoglobulin. In other preferred embodiments, the nucleic acids of interest arc viral nucleic acids, such as components of an adenoviral genome, AAV genome, or retroviral genome. In some embodiments, the present invention allows for co-integration of an expression construct encoding a first subunit of a protein of interest with an expression construct encoding a second subunit of a protein of interest where the expression construct encoding one of the subunits is provided at an increased ratio as compared to the expression construct encoding the other subunits(s). Exemplary ratios are provided below. In some preferred embodiments, the first and second proteins of interest are subunits or components of a viral particle, e.g., the REP and CAP proteins of an AAV vector. In some preferred embodiments, the nucleic acids of interest are viral nucleic acids, such as components of an adenoviral genome, AAV genome, or retroviral genome. In some embodiments, the present invention allows for co-integration of an expression construct encoding a first subunit of a viral particle with an expression construct encoding a second subunit of a viral particle where the expression construct encoding one of the subunits is provided at an increased ratio as compared to the expression construct encoding the other subunits(s). Exemplary ratios are provided below.In some preferred embodiments, the methods comprise introducing at least first nucleic acid constructs encoding a first protein or nucleic acid of interest and second nucleic acid constructs encoding a second protein of interest at a ratio of first nucleic acid constructs to second nucleic acid constructs of at least 1:1, and most preferably at a ratio of from 1:1 to 5000:1 into a host cell having genome comprising from 1 to 500 integrated docking sites, each docking site comprising at least one dock site insertion element and the nucleic acid constructs each comprising at least one insertion element compatible with the at least one dock site insertion element in the integrated docking sites, under conditions such that the nucleic acid expression constructs are inserted at the dock sites at a ratio of first nucleic acid constructs to second nucleic acid constructs of from 1:1 to 1000:1.In some preferred embodiments, the nucleic acid expression constructs are inserted at the dock sites at a ratio of first nucleic acid constructs to second nucleic acid constructs of at least 1.1:1. In some preferred embodiments, the nucleic acid expression constructs are inserted at the dock sites at a ratio of first nucleic acid constructs to second nucleic acid constructs of at least 1.2:1. In some preferred embodiments, the nucleic acid expression constructs are inserted at thedock sites at a ratio of first nucleic acid constructs to second nucleic acid constructs of at least 1.3:1. In some preferred embodiments, the nucleic acid expression constructs arc inserted at the dock sites at a ratio of first nucleic acid constructs to second nucleic acid constructs of at least 1.4:1. In some preferred embodiments, the nucleic acid expression constructs are inserted at the dock sites at a ratio of first nucleic acid constructs to second nucleic acid constructs of at least 1.5:1.In some further preferred embodiments, the ratio of first nucleic acid constructs to second nucleic acid constructs introduced in the host cells to achieve integration is from 1:1 to 5000:1 (including ranges therein, e.g., 1:1 to 4000:1, 1:1 to 3000:1, 1:1 to 2000:1, 1:1 to 1000:1, etc.). In some preferred embodiments, the ratio of first nucleic acid constructs to second nucleic acid constructs introduced in the host cells is from 1.1:1 to 5000:1 (including ranges therein, e.g., 1:1.1 to 4000:1, 1.1:1 to 3000:1, 1:1.1 to 2000:1, 1:1.1 to 1000:1, etc.). In some preferred embodiments, the ratio of first nucleic acid constructs to second nucleic acid constructs is from 1.2:1 to 5000:1 (including ranges therein, e.g., 1:1.2 to 4000:1, 1:1.2 to 3000:1, 1:1.2 to 2000:1, 1:1.2 to 1000:1, etc.). In some preferred embodiments, the ratio of first nucleic acid constructs to second nucleic acid constructs is from 1.3:1 to 5000:1 (including ranges therein, e.g., 1:1.3 to 4000:1, 1:1.3 to 3000:1, 1:1.3 to 2000:1, 1:1.3 to 1000:1, etc.). In some preferred embodiments, the ratio of first nucleic acid constructs to second nucleic acid constructs is from 1.4:1 to 5000:1 (including ranges therein, e.g., 1:1.4 to 4000:1, 1:1.4 to 3000:1, 1:1.4 to 2000:1, 1:1.4 to 1000:1, etc.). In some preferred embodiments, the ratio of first nucleic acid constructs to second nucleic acid constructs is from 1.5:1 to 5000:1 (including ranges therein, e.g., 1:1.5 to 4000:1, 1:1.5 to 3000:1, 1:1.5 to 2000:1, 1:1.5 to 1000:1, etc.). In some preferred embodiments, the ratio of first nucleic acid constructs to second nucleic acid constructs is from 2: 1 to 5000: 1 (including ranges therein, e.g., 2:1 to 4000:1, 2:1 to 3000:1, 2:1 to 2000:1, 2:1 to 1000:1, etc.). In some preferred embodiments, the ratio of first nucleic acid constructs to second nucleic acid constructs is from 5:1 to 5000:1 (including ranges therein, e.g., 5:1 to 4000:1, 5:1 to 3000:1, 5:1 to 2000:1, 5:1 to 1000:1, etc.). In some preferred embodiments, the ratio of first nucleic acid constructs to second nucleic acid constructs is from 10:1 to 5000:1 (including ranges therein, e.g., 10:1 to 4000:1, 10:1 to 3000:1, 10:1 to 2000:1, 10:1 to 1000:1, etc.). In some preferred embodiments, the ratio of first nucleic acid constructs to second nucleic acid constructs is from 20: 1 to 5000: 1 (including ranges therein, e.g., 20:1 to 4000:1, 20:1 to 3000:1, 20:1 to 2000:1, 20:1 to 1000:1, etc.). In somepreferred embodiments, the ratio of first nucleic acid constructs to second nucleic acid constructs is from 10:1 to 200:1. In some preferred embodiments, the ratio of first nucleic acid constructs to second nucleic acid constructs is from 10:1 to 100:1.In still other preferred embodiments, the present invention provides host cells comprising a plurality of docking sites integrated into the genome of the host cell, each docking site comprising at least one dock site insertion element; at least integrated first nucleic acid constructs comprising at least one insertion element compatible with the dock site insertion element and encoding a first protein or nucleic acid of interest, and at least integrated second nucleic acid constructs comprising at least one insertion element compatible with the dock site insertion and encoding a second protein or nucleic acid of interest, wherein the at least integrated first nucleic acid constructs and the at least second integrated nucleic acid constructs are integrated at the plurality of docking sites at a ratio of first nucleic acid constructs to second nucleic acid constructs of at least 1.1:1.In some preferred embodiments, the ratio of integrated first nucleic acid constructs to integrated second nucleic acid constructs introduced in the host cells is from 2:1 to 1000:1. In some preferred embodiments, the ratio of first nucleic acid constructs to second nucleic acid constructs is from 5:1 to 500:1. In some preferred embodiments, the ratio of first nucleic acid constructs to second nucleic acid constructs is from 5:1 to 200:1. In some preferred embodiments, the ratio of first nucleic acid constructs to second nucleic acid constructs is from 10:1 to 200:1. In some preferred embodiments, the ratio of first nucleic acid constructs to second nucleic acid constructs is from 10:1 to 100:1.In some further pref emed embodiments, first, second and third nucleic acid constructs encoding, respectively, first, second and third proteins or nucleic acids of interest are integrated into the host cells at a ratio of from 1:1:1 to 1000:1:1. In some further pref emed embodiments, first, second and third nucleic acid constructs encoding, respectively, first, second and third proteins of interest are integrated into the host cells at a ratio of from 1.5: 1 : 1 to 1000:1:1. In some further preferred embodiments, first, second and third nucleic acid constructs encoding, respectively, first, second and third proteins or nucleic acids of interest are integrated into the host cells at a ratio of from 2:1:1 to 1000:1:1. In some further prefemed embodiments, first, second and third nucleic acid constructs encoding, respectively, first, second and third proteins of interest or nucleic acids are integrated into the host cells at a ratio of from 10: 1 : 1 to 1000: 1:1.In some further preferred embodiments, first, second and third nucleic acid constructs encoding, respectively, first, second and third proteins or nucleic acids of interest arc integrated into the host cells at a ratio of from 1:1:1 to 100:1:1. In some further preferred embodiments, first, second and third nucleic acid constructs encoding, respectively, first, second and third proteins or nucleic acids of interest are integrated into the host cells at a ratio of from 1.5: 1 : 1 to 100:1:1. In some further preferred embodiments, first, second and third nucleic acid constructs encoding, respectively, first, second and third proteins or nucleic acids of interest are integrated into the host cells at a ratio of from 2:1:1 to 100:1:1. In some further preferred embodiments, first, second and third nucleic acid constructs encoding, respectively, first, second and third proteins or nucleic acids of interest are integrated into the host cells at a ratio of from 10:1:1 to 100:1:1.In some further preferred embodiments, first, second and third nucleic acid constructs encoding, respectively, first, second and third proteins or nucleic acids of interest are integrated into the host cells at a ratio of from 1:1:1 to 1000:100:1. In some further preferred embodiments, first, second and third nucleic acid constructs encoding, respectively, first, second and third proteins or nucleic acids of interest are integrated into the host cells at a ratio of from 1.5: 1 : 1 to 1000:100:1. In some further preferred embodiments, first, second and third nucleic acid constructs encoding, respectively, first, second and third proteins or nucleic acids of interest are integrated into the host cells at a ratio of from 2:1:1 to 1000: 100: 1. In some further preferred embodiments, first, second and third nucleic acid constructs encoding, respectively, first, second and third proteins or nucleic acids of interest are integrated into the host cells at a ratio of from 10:1:1 to 1000:100:1.In some further preferred embodiments, first, second and third nucleic acid constructs encoding, respectively, first, second and third proteins or nucleic acids of interest are integrated into the host cells at a ratio of from 1:1:1 to 100:10:1. In some further preferred embodiments, first, second and third nucleic acid constructs encoding, respectively, first, second and third proteins of interest or nucleic acids are integrated into the host cells at a ratio of from 1.5:1:1 to 100:10:1. In some further preferred embodiments, first, second and third nucleic acid constructs encoding, respectively, first, second and third proteins or nucleic acids of interest are integrated into the host cells at a ratio of from 2: 1 : 1 to 100: 10: 1. In some further preferred embodiments, first, second and third nucleic acid constructs encoding, respectively, first, second and thirdproteins of interest or nucleic acids are integrated into the host cells at a ratio of from 10:1 :1 to 100:10:1.In some further preferred embodiments, first, second, third and fourth nucleic acid constructs encoding, respectively, first, second, third and fourth proteins or nucleic acids of interest are integrated into the host cells at a ratio of from 1 : 1 : 1 : 1 to 1000: 1 : 1 : 1. In some further preferred embodiments, first, second, third and fourth nucleic acid constructs encoding, respectively, first, second, third and fourth proteins or nucleic acids of interest are integrated into the host cells at a ratio of from 1.5: 1 : 1 : 1 to 1000: 1 : 1 : 1. In some further preferred embodiments, first, second, third and fourth nucleic acid constructs encoding, respectively, first, second, third and fourth proteins or nucleic acids of interest are integrated into the host cells at a ratio of from 2: 1 : 1 : 1 to 1000: 1 : 1 : 1. In some further preferred embodiments, first, second, third and fourth nucleic acid constructs encoding, respectively, first, second, third and fourth proteins or nucleic acids of interest are integrated into the host cells at a ratio of from 10:1:1:1 to 1000: 1:1:1.In some further preferred embodiments, first, second, third and fourth nucleic acid constructs encoding, respectively, first, second, third and fourth proteins or nucleic acids of interest are integrated into the host cells at a ratio of from 1 : 1 : 1 : 1 to 100: 1 : 1 : 1. In some further preferred embodiments, first, second, third and fourth nucleic acid constructs encoding, respectively, first, second, third and fourth proteins or nucleic acids of interest are integrated into the host cells at a ratio of from 1.5: 1 : 1 : 1 to 100:1:1:1. In some further preferred embodiments, first, second, third and fourth nucleic acid constructs encoding, respectively, first, second, third and fourth proteins or nucleic acids of interest are integrated into the host cells at a ratio of from 2: 1 : 1 : 1 to 100: 1 : 1 : 1. In some further preferred embodiments, first, second, third and fourth nucleic acid constructs encoding, respectively, first, second, third and fourth proteins or nucleic acids of interest are integrated into the host cells at a ratio of from 10:1:1:1 to 100:1:1:1.In some further preferred embodiments, first, second, third and fourth nucleic acid constructs encoding, respectively, first, second, third and fourth proteins or nucleic acids of interest are integrated into the host cells at a ratio of from 1 : 1 : 1 : 1 to 1000: 100: 1 : 1. In some further preferred embodiments, first, second, third and fourth nucleic acid constructs encoding, respectively, first, second, third and fourth proteins or nucleic acids of interest are integrated into the host cells at a ratio of from 1.5: 1 : 1 : 1 to 1000: 100: 1 : 1. In some further preferred embodiments, first, second, third and fourth nucleic acid constructs encoding, respectively, first,second, third and fourth proteins or nucleic acids of interest are integrated into the host cells at a ratio of from 2: 1:1:1 to 1000:100:1:1. In some further preferred embodiments, first, second, third and fourth nucleic acid constructs encoding, respectively, first, second, third and fourth proteins or nucleic acids of interest are integrated into the host cells at a ratio of from 10:1:1:1 to 1000:100:1:1.In some further preferred embodiments, first, second, third and fourth nucleic acid constructs encoding, respectively, first, second, third and fourth proteins or nucleic acids of interest are integrated into the host cells at a ratio of from 1 : 1 : 1 : 1 to 1000: 100: 100: 1. In some further preferred embodiments, first, second, third and fourth nucleic acid constructs encoding, respectively, first, second, third and fourth proteins or nucleic acids of interest are integrated into the host cells at a ratio of from 1.5: 1 : 1 : 1 to 1000:100:100:1. In some further preferred embodiments, first, second, third and fourth nucleic acid constructs encoding, respectively, first, second, third and fourth proteins or nucleic acids of interest are integrated into the host cells at a ratio of from 2: 1:1:1 to 1000:100:100:1. In some further preferred embodiments, first, second, third and fourth nucleic acid constructs encoding, respectively, first, second, third and fourth proteins or nucleic acids of interest are integrated into the host cells at a ratio of from 10:1:1:1 to 1000:100:100:1.In some further preferred embodiments, first, second, third and fourth nucleic acid constructs encoding, respectively, first, second, third and fourth proteins or nucleic acids of interest are integrated into the host cells at a ratio of from 1 : 1 : 1 : 1 to 1000: 100: 100: 1. In some further preferred embodiments, first, second, third and fourth nucleic acid constructs encoding, respectively, first, second, third and fourth proteins or nucleic acids of interest are integrated into the host cells at a ratio of from 1.5: 1:1:1 to 1000:100:100:1. In some further preferred embodiments, first, second, third and fourth nucleic acid constructs encoding, respectively, first, second, third and fourth proteins or nucleic acids of interest are integrated into the host cells at a ratio of from 2: 1:1:1 to 1000:100:100:1. In some further preferred embodiments, first, second, third and fourth nucleic acid constructs encoding, respectively, first, second, third and fourth proteins or nucleic acids of interest are integrated into the host cells at a ratio of from 10:1:1:1 to 1000:100:100:1.In some further preferred embodiments, first, second, third and fourth nucleic acid constructs encoding, respectively, first, second, third and fourth proteins or nucleic acids ofinterest are integrated into the host cells at a ratio of from 1 : 1 : 1 : 1 to 100: 100: 100: 1 . In some further preferred embodiments, first, second, third and fourth nucleic acid constructs encoding, respectively, first, second, third and fourth proteins or nucleic acids of interest are integrated into the host cells at a ratio of from 1.5: 1 : 1 : 1 to 100:100:100:1. In some further preferred embodiments, first, second, third and fourth nucleic acid constructs encoding, respectively, first, second, third and fourth proteins or nucleic acids of interest are integrated into the host cells at a ratio of from 2: 1:1:1 to 100:100:100:1. In some further preferred embodiments, first, second, third and fourth nucleic acid constructs encoding, respectively, first, second, third and fourth proteins or nucleic acids of interest are integrated into the host cells at a ratio of from 10:1:1:1 to 100:100:100:1.In some preferred embodiments, the present invention provides methods for production of a product(s) of interest. In these embodiments, the host cells as described above are cultured in a culture medium under conditions such that the first product of interest and / or two or more products of interest and / or an assembled complex thereof is produced. The product of interest is then isolated or purified from the culture medium. In some embodiments, the product (or products) of interest is operably associated with a signal sequence so the product (or products) are secreted into the culture medium and are then isolated therefrom.In some preferred embodiments, the host cells (and cultures of host cells) are engineered to comprise a plurality of integrated docking sites. For example, in some preferred embodiments, the genomes of the host cells of the present invention preferably comprise from 1 to 1000 integrated docking sites, each docking site comprising at least one dock site insertion element. In other preferred embodiments, the genome of the host cells comprises from 1 to 500 integrated docking sites, each docking site comprising at least one dock site insertion element. In other preferred embodiments, the genome of the host cells comprises from 5 to 500 integrated docking sites, each docking site comprising at least one dock site insertion element. In other preferred embodiments, the genome of the host cells comprises from 5 to 250 integrated docking sites, each docking site comprising at least one dock site insertion element. In other preferred embodiments, the genome of the host cell comprises from 5 to 250 integrated docking sites, each docking site comprising at least one dock site insertion element. In other preferred embodiments, the genome of the host cell comprises from 5 to 100 integrated docking sites, each docking site comprising at least one dock site insertion element. In other preferred embodiments, the genomeof the host cell comprises from 5 to 50 integrated docking sites, each docking site comprising at least one dock site insertion clement. In some preferred embodiments, the integrated docking sites are independent integrated docking sites that are separated from one another and positioned at independent sites within the genome. For example, the integrated docking sites may preferably be spread across a number of chromosome sin the genome. In other embodiments, the integrated docking sites may be present as concatemers which comprise multiple copies of the same DNA sequence linked in series.The integrated docking sites preferably comprise one or more insertion elements (which may be termed a “dock site insertion element.” The dock site insertion elements are preferably nucleic acid sequences that facilitate insertion of a nucleic acid sequence encoding a protein of interest at the dock site. Nucleic acid constructs that can be inserted into the dock sites in the host cells of the present invention are described in detail below.The present invention is not limited to the use of any particular insertion elements. Indeed the use of a variety of insertion elements is contemplated. In some preferred embodiments, the insertion element is a recombinase dock site insertion element. Recombinase dock site insertion elements are nucleic acid sequences that are recognized and utilized by recombinase enzymes.For example, in some preferred embodiments, the recombinase dock site insertion element comprises an attachment site (att). In some particularly preferred embodiments, the attachment site is attP. These attachment sites are utilized by the PhiC31 integrase, which is a recombinase enzyme and which can be provided in the host cell via a vector in preferred embodiments. These dock sites serve as acceptors for integration of nucleic acid constructs comprising an attB attachment site. In other preferred embodiments, attR and attL attachment sites are utilizedIn other preferred embodiments, the recombinase dock site insertion element comprises an Flp Recombination Target (FRT) site. These sites are utilized by the enzyme flippase, which is a recombinase enzyme and which can be provided in the host cell via a vector in preferred embodiments. These dock sites serve as acceptors for integration of nucleic acid constructs comprising at the FRT site.In other preferred embodiments, the recombinase dock site insertion element comprises a LoxP site. These sites are utilized by the Cre recombinase which can be provided in the host cellvia a vector in preferred embodiments. These dock sites serve as acceptors for integration of nucleic acid constructs comprising the LoxP site.In other preferred embodiments, the insertion element is an HDR (homology directed repair) dock site insertion element. HDR dock site insertion elements are nucleic acid sequences that provide an area of homology (a “homology arm”) that base pair with corresponding homology arms on the nucleic acid construct that is inserted at the site. These systems are preferably used with endonucleases that introduce double stranded breaks at a targeted site or sites, preferably flanked by the homology arms. In some embodiments, the HDR dock site insertion element is an AAVS1 safe harbor locus. In these embodiments, the dock site is used utilized by the Rep 78 endonuclease (nickase) which may be introduced into the host cell via a vector. The Rep 78 protein nickase promotes site- specific integration of nucleic acid sequences bearing homology arms corresponding to the AAVS1 safe harbor locus.In other preferred embodiments, the HDR dock site insertion element comprises one or more homology arms that are exogenous sequences of from 30 to 1000 base pairs in length. These dock sites are preferably used in conjunction with CRISPR gene editing systems. In some embodiments, the dock site further comprises one or more sequences that are homologous to guide RNA sequences. In these embodiments, the nucleic acid construct that is inserted at the dock site preferably comprises homology arms that are homologous to and base pair with the homology arms in the dock site. For utilization with CRISPR gene editing systems, a CRISPR gene editing system-compatible nuclease is introduced into the host cell. The CRISPR gene editing system-compatible nuclease may be a wild-type endonuclease that creates a doublestranded break at a position determined by the guide RNA (and within the docking site) or a mutated nuclease (i.e., a nickase) that creates a single stranded break at a staggered positions within the dock site defined by two guide RNAs. Suitable nucleases are described in detail below in the discussion of nucleic acid expression constructs.In some preferred embodiments, the docking site may preferably comprise a suitable promoter so that a promoter trap scheme is utilized when suitable nucleic acid constructs are introduced at the docking site. Suitable promoters include, but are not limited to, SIN-LTR, SV40, EFla, E. coli lac, E. coli trp, phage lambda PL, phage lambda PR, T3, T7, cytomegalovirus (CMV) immediate early, herpes simplex virus (HSV) thymidine kinase, alphalactalbumin, and mouse metallothionein-I promoter sequences. In some preferred embodimentsthe promoter sequence is oriented at the dock site so that the promoter will drive expression from an inserted nucleic acid construct. In some preferred embodiments, the promoter is oriented 5’ to the docking site. In some particularly preferred embodiments, the promoter is a SIN LTR. In these embodiments, the SIN-LTR and EPR are positioned 5’ to the dock site and a SIN LTR is positioned 3’ to the dock site.The docking sites may be introduced into any suitable host cell line. Suitable host cell lines include, but are not limited to, Chinese hamster ovary cells (CH0-K1, ATCC CCL61); bovine mammary epithelial cells (ATCC CRL 10274; bovine mammary epithelial cells); monkey kidney CV1 line transformed by SV40 (COS-7, ATCC CRL 1651); human embryonic kidney line (293 or 293 cells subcloned for growth in suspension culture; see, e.g., Graham et al., J. Gen Virol., 36:59

[1977] ); baby hamster kidney cells (BHK, ATCC CCL 10); mouse sertoli cells (TM4, Mather, Biol. Reprod. 23:243-251

[1980] ); monkey kidney cells (CV1 ATCC CCL 70); African green monkey kidney cells (VERO-76, ATCC CRL- 1587); human cervical carcinoma cells (HELA, ATCC CCL 2); canine kidney cells (MDCK, ATCC CCL 34); buffalo rat liver cells (BRL 3A, ATCC CRL 1442); human lung cells (W138, ATCC CCL 75); human liver cells (Hep G2, HB 8065); mouse mammary tumor (MMT 060562, ATCC CCL51); TRI cells (Mather et al., Annals N.Y. Acad. Sci., 383:44-68

[1982] ); MRC 5 cells; FS4 cells; rat fibroblasts (208F cells); MDBK cells (bovine kidney cells); CAP (CEVEC's Amniocyte Production) cells; VERO cells, NS0 cells, and a human hepatoma line (Hep G2).In some particularly preferred embodiments, the host cells are modified so that they are deficient, or are naturally deficient, in an enzyme activity that is required for growth or survival of the cells in the presence of a selection agent and which is provided by the selectable marker. For example, Chinese Hamster Ovary (CHO) cells have been modified to be deficient for GS . In some preferred embodiments where vector includes a GS selectable marker, the host cell line is deficient in GS. In some particularly preferred embodiments, the GS deficient host cell line is the CHOZN® GS"" cell line available from Merck KGaA. In other embodiments, where the selectable marker is, for example, DHFR, the cell line may preferably be deficient for DHFR activity (i.e., DHFR"). Suitable DHFR- cell lines include but are not limited to CHO-DG44 and derivatives thereof.The docking site sequences may be introduced into the host cells by any suitable genome modification system. In some preferred embodiments, the docking sites are incorporated into thehost cells via the use of integrating vectors. The use of integrating vectors to introduce high copy numbers of a sequence of interest, such as a docking site, is described in detail in US Pat. Nos. 6,852,510 and 7,332,333 as well as US Publ. Nos. 20030092882, 20030224415, 20040235173 and 20050100952, all which are incorporated herein by reference in their entirety.According to the present invention, host cells such as those described above are transduced or transfected with integrating vectors comprising a dock site under conditions such that multiple copies of the dock site are integrated into the genome of the host cell. Examples of integrating vectors include, but are not limited to, retroviral vectors, lentiviral vectors, adeno- associated viral vectors, and transposon vectors.In some preferred embodiments, nucleic acid constructs for expression of a product of interest are introduced into the host cell lines containing multiple docking sites. As discussed above, in preferred embodiments, the nucleic acid constructs preferably comprise nucleic acid sequences (which may be termed “expression construct insertion elements”) that are compatible with the dock site insertion elements as described above.Accordingly, in some preferred embodiments, the present invention provides nucleic acid expression constructs for use in expressing a protein or proteins of interest in a host cell, and in particular to expression of two or more proteins of interest where the nucleic acid expression constructs encoding the two or more proteins of interest are integrated into the genome of the host cell at desired ratios as described in detail above.In some preferred embodiments, where the dock site does not comprise a promoter, the nucleic acid expression constructs, for example, comprise the following elements in operable association, most preferably in 5’ to 3’ order: first promoter sequence - selectable marker sequence - second promoter sequence - nucleic acid sequence encoding a first product of interest - poly A signal sequence. first promoter sequence - selectable marker sequence - second promoter sequence - nucleic acid sequence encoding a second product of interest - poly A signal sequence. first promoter sequence - selectable marker sequence - second promoter sequence - nucleic acid sequence encoding a third product of interest - poly A signal sequence.first promoter sequence - selectable marker sequence - second promoter sequence - nucleic acid sequence encoding a fourth product of interest - poly A signal sequence.In some preferred embodiments, where the dock site comprises an exogenous promoter, the nucleic acid expression constructs, for example, comprise the following elements in operable association, most preferably in 5’ to 3’ order: selectable marker sequence - second promoter sequence - nucleic acid sequence encoding a first product of interest - poly A signal sequence. selectable marker sequence - second promoter sequence - nucleic acid sequence encoding a second product of interest - poly A signal sequence. selectable marker sequence - second promoter sequence - nucleic acid sequence encoding a third product of interest - poly A signal sequence. selectable marker sequence - second promoter sequence - nucleic acid sequence encoding a fourth product of interest - poly A signal sequence.It will be understood that the invention is not limited to expression of any particular number of products of interest and that the exemplary sets of constructs described above and below may be adapted for the expression of 2, 3, 4, 5, 6, 7, 8, 8, 10, etc. products of interest.In some preferred embodiments, the constructs of the invention do not comprise a poly A signal sequence between the selectable marker sequence and second promoter sequence. The present invention is not limited to any particular mechanism of action. Indeed, an understanding of the mechanism of action is not necessary to practice the present invention. Nevertheless, constructs which lack a poly A signal sequence after the selectable marker have been found to provide for better selection and production of the protein of interest in host cell cultures. In still other preferred embodiments, the selectable marker is adjacent to the second promoter. In still other preferred embodiments, the second promoter is adjacent to the nucleic acid sequenceencoding the first protein of interest. In this context, the term “adjacent” means that there is no intervening functional element or intron between the listed components.In some particularly preferred embodiments, the nucleic acid expression constructs further comprises at least one expression construct insertion element at a position or positions selected from the group consisting of 5’ to the first promoter, 3’ to the poly A signal sequence, between the first promoter and the poly A signal sequence, between the selectable marker and the second promoter sequence, and both 5’ to the first promoter and 3’ to the poly A signal sequence. Suitable constructs are shown in the following non-limiting examples: expression construct insertion element - first promoter sequence (optional depending on whether the dock site already comprises an exogenous promoter sequence) - selectable marker sequence - second (i.e., internal) promoter sequence - nucleic acid sequence encoding a first product of interest - poly A signal sequence first promoter sequence (optional depending on whether the dock site already comprises an exogenous promoter sequence) - selectable marker sequence - second promoter sequence - nucleic acid sequence encoding a first product of interest - poly A signal sequence - expression construct insertion element expression construct insertion element - first promoter sequence (optional depending on whether the dock site already comprises an exogenous promoter sequence) - selectable marker sequence - second promoter sequence - nucleic acid sequence encoding a first product of interest - poly A signal sequence - expression construct insertion element. first promoter sequence (optional depending on whether the dock site already comprises an exogenous promoter sequence)- selectable marker sequence - expression construct insertion element - second promoter sequence - nucleic acid sequence encoding a first product of interest - poly A signal sequence. expression construct insertion element - first promoter sequence (optional depending on whether the dock site already comprises an exogenous promoter sequence) - selectablemarker sequence - second (i.e., internal) promoter sequence - nucleic acid sequence encoding a second product of interest - poly A signal sequence first promoter sequence (optional depending on whether the dock site already comprises an exogenous promoter sequence) - selectable marker sequence - second promoter sequence - nucleic acid sequence encoding a second product of interest - poly A signal sequence - expression construct insertion element expression construct insertion element - first promoter sequence (optional depending on whether the dock site already comprises an exogenous promoter sequence) - selectable marker sequence - second promoter sequence - nucleic acid sequence encoding a second product of interest - poly A signal sequence - expression construct insertion element. first promoter sequence (optional depending on whether the dock site already comprises an exogenous promoter sequence)- selectable marker sequence - expression construct insertion element - second promoter sequence - nucleic acid sequence encoding a second product of interest - poly A signal sequence.In some preferred embodiments, the constructs may include nucleic acid sequences encoding multiple products of interest, for example 2, 3 ,4 or 5 (or more) products of interest. Suitable constructs for expressing two products of interest are shown in the following nonlimiting examples. These expression constructs may be used at different ratios in conjunction with expression constructs encoding an additional third product of interest, or as exemplified below, third and fourth products of interest. expression construct insertion element - first promoter sequence (optional depending on whether the dock site already comprises an exogenous promoter sequence) - selectable marker sequence - second (i.e., internal) promoter sequence - nucleic acid sequence encoding a first product of interest - WPRE (optional) - poly A signal sequence - third promoter sequence or IRES - nucleic acid sequence encoding a second product of interest - WPRE (optional) - poly A signal sequencefirst promoter sequence (optional depending on whether the dock site already comprises an exogenous promoter sequence) - selectable marker sequence - second promoter sequence - nucleic acid sequence encoding a first product of interest - WPRE (optional)- poly A signal sequence - third promoter sequence - intron (optional) - nucleic acid sequence encoding a second product of interest - WPRE (optional) - poly A signal sequence - expression construct insertion element expression construct insertion element - first promoter sequence (optional depending on whether the dock site already comprises an exogenous promoter sequence) - selectable marker sequence - second promoter sequence - nucleic acid sequence encoding a first product of interest - WPRE (optional) - poly A signal sequence - third promoter sequence - intron (optional) - nucleic acid sequence encoding a second product of interest - WPRE (optional) - poly A signal sequence - expression construct insertion element. first promoter sequence (optional depending on whether the dock site already comprises an exogenous promoter sequence) - selectable marker sequence - expression construct insertion element - second promoter sequence - nucleic acid sequence encoding a first product of interest - WPRE - poly A signal sequence - third promoter sequence or IRES- nucleic acid sequence encoding a second product of interest - WPRE - poly A signal sequence expression construct insertion element - first promoter sequence (optional depending on whether the dock site already comprises an exogenous promoter sequence) - selectable marker sequence - second promoter sequence - nucleic acid sequence encoding a first product of interest - WPRE (optional) - poly A signal sequence - third promoter sequence - nucleic acid sequence encoding a second product of interest - WPRE (optional) - poly A signal sequence - expression construct insertion element.expression construct insertion element - first promoter sequence (optional depending on whether the dock site already comprises an exogenous promoter sequence) - selectable marker sequence - second promoter sequence - nucleic acid sequence encoding a first product of interest - WPRE (optional) - poly A signal sequence - third promoter sequence - intron- nucleic acid sequence encoding a second product of interest - WPRE (optional) - poly A signal sequence - expression construct insertion element. expression construct insertion element - first promoter sequence (optional depending on whether the dock site already comprises an exogenous promoter sequence) - selectable marker sequence - second (i.e., internal) promoter sequence - nucleic acid sequence encoding a third product of interest - WPRE (optional) - poly A signal sequence - third promoter sequence or IRES - nucleic acid sequence encoding a fourth product of interest- WPRE (optional) - poly A signal sequence first promoter sequence (optional depending on whether the dock site already comprises an exogenous promoter sequence) - selectable marker sequence - second promoter sequence - nucleic acid sequence encoding a third product of interest - WPRE (optional)- poly A signal sequence - third promoter sequence - intron (optional) - nucleic acid sequence encoding a fourth product of interest - WPRE (optional) - poly A signal sequence - expression construct insertion element expression construct insertion element - first promoter sequence (optional depending on whether the dock site already comprises an exogenous promoter sequence) - selectable marker sequence - second promoter sequence - nucleic acid sequence encoding a third product of interest - WPRE (optional) - poly A signal sequence - third promoter sequence - intron (optional) - nucleic acid sequence encoding a fourth product of interest - WPRE (optional) - poly A signal sequence - expression construct insertion element. first promoter sequence (optional depending on whether the dock site already comprises an exogenous promoter sequence) - selectable marker sequence - expression constructinsertion element - second promoter sequence - nucleic acid sequence encoding a third product of interest - WPRE - poly A signal sequence - third promoter sequence or IRES - nucleic acid sequence encoding a fourth product of interest - WPRE - poly A signal sequence expression construct insertion element - first promoter sequence (optional depending on whether the dock site already comprises an exogenous promoter sequence) - selectable marker sequence - second promoter sequence - nucleic acid sequence encoding a third product of interest - WPRE (optional) - poly A signal sequence - third promoter sequence - nucleic acid sequence encoding a fourth product of interest - WPRE (optional) - poly A signal sequence - expression construct insertion element. expression construct insertion element - first promoter sequence (optional depending on whether the dock site already comprises an exogenous promoter sequence) - selectable marker sequence - second promoter sequence - nucleic acid sequence encoding a third product of interest - WPRE (optional) - poly A signal sequence - third promoter sequence - intron- nucleic acid sequence encoding a fourth product of interest - WPRE (optional) - poly A signal sequence - expression construct insertion element.In some embodiments, mixtures of different constructs are utilized. In some preferred embodiments, the mixture of different constructs may comprise constructs as described above and constructs starting with the internal or second promoter (i.e., starting after and not including the selectable marker). It is contemplated that by using mixtures of constructs, some which do not include selectable markers, that higher insertion rates may be achieved.However, any suitable products of interest may be expressed via the host cells, constructs and systems of the present invention. Exemplary products of interest include immunoglobulins, single chain antibodies, anticoagulant proteins, blood factor proteins, bone morphogenetic proteins, engineered protein scaffolds, enzymes, Fc fusion proteins, growth factors, hormones, interferons, interleukins, antigens, and thrombolytic proteins.In other preferred embodiments, the constructs of the present invention may be utilized to express viral vector sequences as the product of interest. Viral vector expression sequences thatmay be included in the constructs of the present invention include, but are not limited to, retroviral vectors, lentiviral vectors, adenoviral vectors and AAV vectors as described elsewhere herein In some preferred embodiments, the retroviral vectors themselves include a nucleic acid sequence encoding a protein of interest as described above that is expressed by the vector. In some particularly preferred embodiments, the protein of interest that is expressed by the vector is an antigen sequence for use in a vaccine.In some preferred embodiments, the expression construct insertion elements are elements that find use in conjunction with or are recognized by transposons, integrases, recombinases or CRISPR systems. Suitable insertion elements include, but are not limited to, inverted terminal repeats, integrase attachment sites (att), and homologous recombination arms which in the context of the constructs described herein can be described as homologous recombination insertion elements.The nucleic acid constructs may be utilized with many different vectors and vectors systems. These vectors and vectors system may preferably be used to introduce the nucleic acid expression constructs into the host cells described above. Suitable vectors and vectors systems include, but are not limited to, viral gene insertion technologies such as retroviral, lentiviral and AAV systems as well as non-viral gene insertion technologies such as transposase, recombinase, integrase or CRISPR gene insertion. Specific examples of technologies / enzymes that can be used with nucleic acid constructs of the present invention include piggyback transposase systems, sleeping beauty transposase systems, Most transposase systems, Tol2 transposase systems, Leapin transposase systems, Lambda recombinase systems, FLP / FRT systems, Cre / Lox systems, MMLV integrase systems, Rep 78 integrase systems and CRISPR systems which can include nucleases or nickases as well as guide sequences. In some preferred embodiments, the system is a nucleic acid integration system with the proviso that the system is not a retroviral or lentiviral systems utilizing a retroviral or lentiviral LTR.As discussed above, in some preferred embodiments, the expression construct insertion element comprises an attachment site (att). In some particular’ preferred embodiments, the attachment site is attB. These attachment sites are utilized by the PhiC31 integrase, which is a recombinase enzyme and which can be provided in the host cell via a vector in preferred embodiments. These sites facilitate integration of the nucleic acid constructs into a dock sitecomprising attP attachment site. In other preferred embodiments, attR and attL attachment sites may be utilized.In other preferred embodiments, the expression construct insertion element comprises an Flp Recombination Target (FRT) site. These sites are utilized by the enzyme flippase, which is a recombinase enzyme and which can be provided in the host cell via a vector in preferred embodiments. These sites serve facilitate integration of nucleic acid constructs into dock sites comprising corresponding FRT sites.In other preferred embodiments, the expression construct insertion element comprises a LoxP site. These sites are utilized by the Cre recombinase which can be provided in the host cell via a vector in preferred embodiments. These sites facilitate integration of nucleic acid constructs into dock sites comprising corresponding LoxP sites.In other preferred embodiments, the expression construct insertion element is an HDR (homology directed repair) expression construct insertion element. HDR expression construct insertion elements are nucleic acid sequences that provide an area of homology (a “homology arm”) that base pair with corresponding homology arms in the dock site. These systems are preferably used with endonucleases that introduce double stranded breaks at a targeted site or sites, preferably flanked by the homology arms. In some embodiments, the HDR expression construct insertion element comprises AAVS1 safe harbor locus homology arms. In these embodiments, the expression construct is specifically integrated in a dock site comprising the AAVS1 safe harbor locus. The integration is facilitated by the Rep 78 endonuclease (nickase) which may be introduced into the host cell via a vector. The Rep 78 protein nickase promotes site- specific integration of nucleic acid sequences bearing homology arms corresponding to the AAVS1 safe harbor locus.In other preferred embodiments, the HDR expression construct insertion element comprises one or more homology arms that are exogenous sequences of from 30 to 1000 base pairs in length. These expression constructs are preferably used in conjunction with CRISPR gene editing systems. In these embodiments, the nucleic acid construct is inserted at dock sites that comprise homology arms that are homologous to and base pair with the homology arms in the nucleic acid construct. For utilization with CRISPR gene editing systems, a CRISPR gene editing system-compatible nuclease is introduced into the host cell. The CRISPR gene editing system-compatible nuclease may be a wild-type endonuclease that creates a double-strandedbreak at a position determined by the guide RNA (and within the docking site) or a mutated nuclease (i.c., a nickase) that creates a single stranded break at a staggered positions within the dock site defined by two guide RNAs. Suitable nucleases are described in detail below in the discussion of nucleic acid expression constructs.As discussed above, integration at the dock sites generally requires expression of an exogenous enzyme in the host cell. Suitable enzymes include, but are not limited to, recombinases (including integrases), endonucleases, and nickases. Accordingly, in some embodiments, host cells of the present invention comprise an exogenous nucleic acid sequence (or expression construct) for expression of a recombinase (including integrases), an endonuclease, and a nickase In some embodiments, constructs for expressing the exogenous enzymes may be stably integrated into the genome of the host cell. In other embodiments, vectors for expressing the exogenous enzymes are transiently introduced into the host cell, for example with an extrachromosomal vector such as a plasmid.In some embodiments, both the vectors comprising exogenous enzyme and the vectors comprising the nucleic acid constructs for expression of the protein of interest are transiently introduced into the host cell, for example by transfection. In these embodiments, the preferred ratio of the vectors encoding the exogenous enzyme to the gene of interest vectors is from 1:1000 to 1:10. In some more preferred embodiments, the ratio is from 1:100 to 1:750. In some still more preferred embodiments, the ratio is from 1:400 to 1:600. This is surprising as the literature for other integrase systems generally indicates that a higher level of vector encoding the exogenous enzyme to the gene of interest construct is required.In some preferred embodiments, the integrase is the phiC31 integrase (BioCat GmbH, Heidelberg, DE or System Biosciences, Palo Alto, CA)). The phiC31 integrase is a sequencespecific recombinase encoded within the genome of the bacteriophage phiC31. The phiC31 integrase mediates recombination between two 34 base pair sequences termed attachment sites (att), one found in the phage and the other in the host. This serine integrase has been shown to function efficiently in many different cell types including mammalian cells. In the presence of phiC31 integrase, an attB- containing donor plasmid can be unidirectional integrated into a target genome through recombination at sites with sequence similarity to the native attP site (termed pseudo-attP sites). phiC31 integrase can integrate a plasmid of any size, as a single copy, and requires no cofactors. The integrated transgenes are stably expressed and heritable.Other suitable recombinase-based systems include CRISPR gene editing systems, CRE- Eox, FLP-FRT, and lambda recombinase systems.Cre-Eox recombination is a site-specific recombinase technology, used to carry out deletions, insertions, translocations and inversions at specific sites in the DNA of cells. It allows the DNA modification to be targeted to a specific cell type or be triggered by a specific external stimulus. It is implemented both in eukaryotic and prokaryotic systems. The Cre-lox recombination system has been particularly useful to help neuroscientists to study the brain in which complex cell types and neural circuits come together to generate cognition and behaviors. The system consists of a single enzyme, Cre recombinase, that recombines a pair of short target sequences called the Fox sequences. This system can be implemented without inserting any extra supporting proteins or sequences. The Cre enzyme and the original Fox site called the FoxP sequence are derived from bacteriophage PE See, e.g., Targeted integration of DNA using mutant lox sites in embryonic stem cells. Araki, et al. Nucleic Acids Res, Feb 1997, Vol. 25, Issue 4, pp. 868-872; High-Resolution Eabeling and Functional Manipulation of Specific Neuron Types in Mouse Brain by Cre-Activated Viral Gene Expression. Kuhlman, et al. PEos One, Apr 2008, Vol. 3, e2005; When reverse genetics meets physiology: the use of site-specific recombinases in mice. Tranche, et al. FEBS Fetters, Aug 2002, Vol. 529, Issue 1, pp. 116-121.The FEP-FRT recombination system is another site-directed recombination technology very conceptually similar to Cre-lox, with flippase (Flp) and the short flippase recognition target (FRT) site being analogous to Cre and loxP, respectively. See, e.g., Candice et al., Cre / loxP, Flp / FRT Systems and Pluripotent Stem Cell Fines (2012) Topics in Current Genetics, vol 23. The FEP-FRT technology can be an effective alternative to Cre-lox, and has also been used in conjunction with it, allowing for two separate recombination events to be controlled in parallel.The nucleic acid constructs of the present invention may be used in conjunction with CRISPR homologous recombination (HDR) systems. HDR is initiated by the presence of double strand breaks (DSBs) in DNA. The CRISPR / Cas9 system is preferably used to create targeted double stranded breaks via a guide RNA sequence so that the nucleic acid construct of the invention can be inserted. See, e.g., Zhang et al., Efficient precise knockin with a double cut HDR donor after CRISPR / Cas9-mediated double- stranded DNA cleavage (2017) Genome Biol. 18:35; Mali et al., Cas9 as a versatile tool for engineering biology. Nature MethodslO, 957-963 (2013); Mali et al., RNA-Guided Human Genome Engineering via Cas9. Science339(6121), 823-826 (2013); Ran et al., Double nicking by RNA-guided CRISPR Cas9 for enhanced genome editing specificity. Cell, 155(2), 479-480(2013). Suitable guide RNA sequences (gRNAs) may be designed as is known in the art. In some preferred embodiments, CRISPR systems for HDR utilize either one or two guide sequences. When one guide RNA sequence is utilized, it preferred to use a nuclease such as a Cas9 nuclease which makes a single double stranded break guided by the guide RNA sequence. When two guide sequences are utilized, it is preferred to use a nickase, which can be a mutated Cas9 nuclease which only makes single stranded breaks in the target DNA sequence guided by each of the guide RNA sequences. The single stranded breaks are preferably positioned at staggered points on different strands (i.e., the sense and antisense strands) of the target DNA sequence. This arrangement generally improves HDR efficiency.In general, “CRISPR system” refers collectively to transcripts and other elements involved in the expression of or directing the activity of CRISPR-associated (“Cas”) genes, including sequences encoding a Cas gene, a tracr (trans-activating CRISPR) sequence (e.g. tracrRNA or an active partial tracrRNA), a tracr-mate sequence (encompassing a “direct repeat” and a tracrRNA-processed partial direct repeat in the context of an endogenous CRISPR system), a guide sequence (also referred to as a “spacer” in the context of an endogenous CRISPR system), or other sequences and transcripts from a CRISPR locus. In some embodiments, one or more elements of a CRISPR system is derived from a type I, type II, or type III CRISPR system. In some embodiments, one or more elements of a CRISPR system is derived from a particular' organism comprising an endogenous CRISPR system, such as Streptococcus pyogenes. In general, a CRISPR system is characterized by elements that promote the formation of a CRISPR complex at the site of a target sequence (also referred to as a protospacer in the context of an endogenous CRISPR system). In the context of formation of a CRISPR complex, “target sequence” refers to a sequence to which a guide sequence is designed to have complementarity, where hybridization between a target sequence and a guide sequence promotes the formation of a CRISPR complex. Full complementarity is not necessarily required, provided there is sufficient complementarity to cause hybridization and promote formation of a CRISPR complex. A target sequence may comprise any polynucleotide, such as DNA or RNA polynucleotides. In some embodiments, a target sequence is located in the nucleus or cytoplasm of a cell. In some embodiments, the target sequence may be within an organelle of a eukaryotic cell, for example, mitochondrion or chloroplast. A sequence or template that may be used for recombination intothe targeted locus comprising the target sequences is referred to as an “editing template” or “editing polynucleotide” or “editing sequence”. In aspects of the invention, an exogenous template polynucleotide may be referred to as an editing template. In an aspect of the invention the recombination is homologous recombination.Typically, in the context of an endogenous CRISPR system, formation of a CRISPR complex (comprising a guide sequence hybridized to a target sequence and complexed with one or more Cas proteins) results in cleavage of one or both strands in or near (e.g. within 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 50, or more base pairs from) the target sequence. Without wishing to be bound by theory, the tracr sequence, which may comprise or consist of all or a portion of a wildtype tracr sequence (e.g. about or more than about 20, 26, 32, 45, 48, 54, 63, 67, 85, or more nucleotides of a wild-type tracr sequence), may also form pail of a CRISPR complex, such as by hybridization along at least a portion of the tracr sequence to all or a portion of a tracr mate sequence that is operably linked to the guide sequence. In some embodiments, the tracr sequence has sufficient complementarity to a tracr mate sequence to hybridize and participate in formation of a CRISPR complex. As with the target sequence, it is believed that complete complementarity is not needed, provided there is sufficient to be functional. In some embodiments, the tracr sequence has at least 50%, 60%, 70%, 80%, 90%, 95% or 99% of sequence complementarity along the length of the tracr mate sequence when optimally aligned. In some embodiments, one or more vectors driving expression of one or more elements of a CRISPR system are introduced into a host cell such that expression of the elements of the CRISPR system direct formation of a CRISPR complex at one or more target sites. For example, a Cas enzyme, a guide sequence linked to a tracr-mate sequence, and a tracr sequence could each be operably linked to separate regulatory elements on separate vectors. Alternatively, two or more of the elements expressed from the same or different regulatory elements, may be combined in a single vector, with one or more additional vectors providing any components of the CRISPR system not included in the first vector. CRISPR system elements that are combined in a single vector may be arranged in any suitable orientation, such as one element located 5' with respect to (“upstream” of) or 3' with respect to (“downstream” of) a second element. The coding sequence of one element may be located on the same or opposite strand of the coding sequence of a second element, and oriented in the same or opposite direction. In some embodiments, a single promoter drives expression of a transcript encoding a CRISPR enzyme and one or more of the guide sequence, tracr matesequence (optionally operably linked to the guide sequence), and a tracr sequence embedded within one or more intron sequences (c.g. each in a different intron, two or more in at least one intron, or all in a single intron). In some embodiments, the CRISPR enzyme, guide sequence, tracr mate sequence, and tracr sequence are operably linked to and expressed from the same promoter.Non-limiting examples of Cas proteins useful in the present invention include Casl, CaslB, Cas2, Cas3, Cas4, Cas5, Cas6, Cas7, Cas8, Cas9 (also known as Csnl and Csxl2), CaslO, Csyl, Csy2, Csy3, Csel, Cse2, Cscl, Csc2, Csa5, Csn2, Csm2, Csm3, Csm4, Csm5, Csm6, Cmrl, Cmr3, Cmr4, Cmr5, Cmr6, Csbl, Csb2, Csb3, Csxl7, Csxl4, CsxlO, Csxl6, CsaX, Csx3, Csxl, Csxl5, Csfl, Csf2, Csf3, Csf4, homologs thereof, or modified versions thereof. These enzymes are known; for example, the amino acid sequence of 5. pyogenes Cas9 protein may be found in the SwissProt database under accession number Q99ZW2. In some embodiments, the unmodified CRISPR enzyme has DNA cleavage activity, such as Cas9. In some embodiments the CRISPR enzyme is Cas9, and may be Cas9 from .S', pyogenes or 5. pneumoniae. In some embodiments, the CRISPR enzyme directs cleavage of one or both strands at the location of a target sequence, such as within the target sequence and / or within the complement of the target sequence. In some embodiments, the CRISPR enzyme directs cleavage of one or both strands within about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 25, 50, 100, 200, 500, or more base pairs from the first or last nucleotide of a target sequence. In some embodiments, a vector encodes a CRISPR enzyme that is mutated to with respect to a corresponding wild-type enzyme such that the mutated CRISPR enzyme lacks the ability to cleave one or both strands of a target polynucleotide containing a target sequence. For example, an aspartate-to-alanine substitution (D10A) in the RuvC I catalytic domain of Cas9 from S. pyogenes converts Cas9 from a nuclease that cleaves both strands to a nickase (cleaves a single strand). Other examples of mutations that render Cas9 a nickase include, without limitation, H840A, N854A, and N863A. In aspects of the invention, nickases may be used for genome editing via homologous recombination.In some preferred embodiments, the HDR insertion element comprises AAVS1 safe harbor locus homology arms and are used in conjunction with Rep 78 endonuclease (nickase). The adeno-associated virus serotype 2 (AAV2) Rep 78 protein is a strand- specific endonuclease (nickase) that promotes site-specific integration of transgene sequences bearing homology armscorresponding to the AAVS1 safe harbor locus. See, e.g., Ramachandra et al., Efficient rccombinasc-mcdiatcd cassette exchange at the AAVS1 locus in human embryonic stem cells using baculoviral vectors (2011) Nucleic Acids Research, 39(16):el07; WO1998027207).As indicated above, in some preferred embodiments, the nucleic acid constructs of the present invention comprise an optional first and a second promoter sequence. The first and second promoter sequences may be the same or different. Suitable first and second promoter sequences include, but are not limited to the MMLV LTR promoter, the MoMuSV LTR promoter, the RSV LTR promoter, the SIN LTR promoter, the SV40 promoter, cytomegalovirus (CMV) immediate early promoter, herpes simplex virus (HSV) thymidine kinase promoter, alpha-lactalbumin promoter, mouse metallothionein-I promoter, dihydrofolate reductase promoter, the p-actin promoter, phosphoglycerol kinase (PGK) promoter, and the EFla promoter sequences, and combinations thereof. In some preferred embodiments, the first promoter sequence is not a retroviral LTR promoter, i.e.. the first promoter is promoter sequence other than a retroviral LTR promoter sequence. However, when the promoter is a retroviral promoter sequence, it may be a SIN (self-inactivating) LTR promoter sequence. See, e.g., co-pending application PCT / US2019 / 064423, which is incorporated herein by reference in its entirety. Suitable Sin LTR promotors are known in the ail and are prepared by removing either all or a portion of the U3 region of the LTR.As described in PCT / US2019 / 064423, in some preferred embodiments the first promoter which drives selectable marker is a weak promoter. In some preferred embodiments, a weak promoter is a promoter, preferably a constitutive promoter, that has activity that equal to or less than the activity of the SIN LTR promoter in a host of interest (e.g., a CHO cell) when operably linked to a selectable maker sequence. In still other preferred embodiments, a weak promoter is a promoter, preferably a constitutive promoter, that has activity that equal to or less than the activity of the human Ubiquitin C (UBC) promoter in a host of interest (e.g., a CHO cell) when operably linked to a selectable maker sequence. Suitable methods for assessing promoter strength are known in the art. See, e.g., Dandindorj et al. (2014) A Comparative Analysis of Constitutive Promoters Located in Adeno-Associated Viral Vectors, PLoS One 9(8): el06472; Zhang and Baum (2005) Evaluation of Viral and Mammalian Promoters for Use in Gene Delivery to Salivary Glands Mol. Ther. 12(3):528-536; Qin et al. (2010) Systematic Comparison of Constitutive Promoters and the Doxycycline-Inducible Promoter PLoS 5(5): el 0611;Jeyaseelan et al. (2001) Real-time detection of gene promoter activity: quantitation of toxin gene transcription, Nucleic Acids Research. 29 (12): 58c-58. In some embodiments, weak promoters have been altered to reduce promoter activity. Accordingly, in some preferred embodiments, the present invention provides vector(s) for expression of a protein of interest comprising a nucleic acid sequence encoding a selectable marker in operable association with a first weak promoter sequence or promoter sequence that has been altered to reduce promoter activity as compared to a non-altered or wild-type version of the first promoter sequence and a nucleic acid sequence encoding the protein of interest operably linked to a second promoter sequence. The SIN LTR promoter sequence is one such example. Other promoter sequences described above may also be altered to reduce activity and provide a weak promoter or the weak promoter may be naturally occurring weak promoter such as the UBC promoter.In some preferred embodiments, the nucleic acid constructs include a selectable marker. Suitable selectable markers include but are not limited to glutamine synthetase (GS), dihydrofolate reductase (DHFR) and the like. These genes are described in U.S. Pat. Nos. 5,770,359; 5,827,739; 4,399,216; 4,634,665; 5,149,636; and 6,455,275; all of which are incorporated herein by reference. In some preferred embodiments, the selectable marker that is utilized is compatible with a host cell line that is deficient in the production of the enzyme encoded by the selectable marker nucleic acid sequence. Suitable host cell lines are described in more detail below. In other embodiments, the selectable marker is an antibiotic resistance marker, i.e., a gene that produces a protein that provides cells expressing this protein with resistance to an antibiotic. Suitable antibiotic resistance markers include genes that provide resistance to neomycin (neomycin resistance gene (neo)), hygromycin (hygromycin B phosphotransferase gene), puromycin (puromycin N-acetyl-transferase), and the like.In other embodiments of the present invention, where secretion of the protein of interest is desired, the nucleic acid constructs include a signal peptide sequence in operable association with the protein of interest. The sequences of several suitable signal peptides are known to those in the ail, including, but not limited to, those derived from tissue plasminogen activator, human growth hormone, lactoferrin, alpha-casein, and alpha-lactalbumin.In other embodiments of the present invention, the nucleic acid constructs include an RNA export element (See, e.g., U.S. Pat. Nos. 5,914,267; 6,136,597; and 5,686,120; and WO99 / 14310, all of which are incorporated herein by reference) either 3' or 5' to the nucleic acidsequence encoding the protein of interest. Tt is contemplated that the use of RNA export elements allows high levels of expression of the protein of interest without incorporating splice signals or introns in the nucleic acid sequence encoding the protein of interest.In still other embodiments, the nucleic acid constructs include at least one internal ribosome entry site (IRES) sequence. The sequences of several suitable IRES's are available, including, but not limited to, those derived from foot and mouth disease virus (FDV), encephalomyocarditis virus, and poliovirus. The IRES sequence can be interposed between two transcriptional units (e.g., nucleic acids encoding different proteins of interest or subunits of a multi-subunit protein such as an antibody) to form a polycistronic sequence so that the two transcriptional units are transcribed from the same promoter.In some preferred embodiments, the nucleic acid constructs are incorporated into a nucleic acid expression vector. Vectors include, but are not limited to, nucleic acid molecules that are single-stranded, double- stranded, or partially double-stranded; nucleic acid molecules that comprise one or more free ends, no free ends (e.g. circular); nucleic acid molecules that comprise DNA, RNA, or both; and other varieties of polynucleotides known in the art. One type of vector is a “plasmid,” which refers to a circular double stranded DNA loop into which additional DNA segments can be inserted, such as by standard molecular cloning techniques. Another type of vector is a viral vector, wherein virally-derived DNA or RNA sequences are present in the vector for packaging into a virus (e.g. retroviruses, replication defective retroviruses, adenoviruses, replication defective adenoviruses, and adeno-associated viruses). Viral vectors also include polynucleotides carried by a virus for transfection into a host cell. Certain vectors are capable of autonomous replication in a host cell into which they are introduced (e.g. bacterial vectors having a bacterial origin of replication and episomal mammalian vectors). Other vectors (e.g., non-episomal mammalian vectors) are integrated into the genome of a host cell upon introduction into the host cell, and thereby are replicated along with the host genome. Moreover, certain vectors are capable of directing the expression of genes to which they are operatively-linked. Such vectors are referred to herein as “expression vectors.” Common expression vectors of utility in recombinant DNA techniques are often in the form of plasmids. Other suitable vectors include, but are not limited to, cosmids and Yeast Artificial Chromosomes.Accordingly, suitable nucleic acid expression vectors include, but are not limited to, transposon vectors as described above, as well as plasmid vectors, retroviral vectors, lentiviral vectors, AAV vectors, phage vectors, etc.). It is contemplated that any vector may be used as long as it is replicable and viable in the host. In preferred embodiments, the vectors are mammalian expression vectors that comprise among other elements described herein an origin of replication, a suitable promoter and enhancer, and also any necessary ribosome binding sites, polyadenylation sites, splice donor and acceptor sites, transcriptional termination sequences, and 5' flanking non-transcribed sequences.Suitable plasmid vectors that may be adapted to incorporate the nucleic acid constructs of the present invention include specific plasmids systems for transposon vectors, FLP-FLT systems, Cre-lox systems, CRISPR-Cas9 systems, recombinase systems and integrase systems as well as plasmid vectors derived from pCIneo, pVAXl, pACT, Gateway plasmids, pAdvantage, pBIND, pG51uc, pTNT, pTarget, pCat3, pSI, pCMV, pSV and the like.In some embodiments, the present invention provides host cells and host cell culture wherein the host cells express the protein of interest from the nucleic acid constructs described above. In preferred embodiment, the host cells a mammalian host cells. A number of mammalian host cell lines are known in the art. In general, these host cells are capable of growth and survival when placed in either monolayer culture or in suspension culture in a medium containing the appropriate nutrients and growth factors, as is described in more detail below. Typically, the cells are capable of expressing and secreting large quantities of a particular protein of interest into the culture medium. Examples of suitable mammalian host cells include, but are not limited to Chinese hamster ovary cells (CH0-K1, ATCC CC1-61); bovine mammary epithelial cells (ATCC CRL 10274; bovine mammary epithelial cells); monkey kidney CV1 line transformed by SV40 (COS-7, ATCC CRL 1651); human embryonic kidney line (293 or 293 cells subcloned for growth in suspension culture; see, e.g., Graham et al., J. Gen Virol., 36:59

[1977] ); baby hamster kidney cells (BHK, ATCC CCL 10); mouse sertoli cells (TM4, Mather, Biol. Reprod. 23:243-251

[1980] ); monkey kidney cells (CV1 ATCC CCL 70); African green monkey kidney cells (VERO-76, ATCC CRL- 1587); human cervical carcinoma cells (HELA, ATCC CCL 2); canine kidney cells (MDCK, ATCC CCL 34); buffalo rat liver cells (BRL 3 A, ATCC CRL 1442); human lung cells (W138, ATCC CCL 75); human liver cells (Hep G2, HB 8065); mouse mammary tumor (MMT 060562, ATCC CCL51); TRI cells (Mather et al., AnnalsN.Y. Acad. Sci., 383:44-68

[1982] ); MRC 5 cells; FS4 cells; rat fibroblasts (208F cells); MDBK cells (bovine kidney cells); CAP (CEVEC's Amniocytc Production) cells; VERO cells, NSO cells, and a human hepatoma line (Hep G2).In some particularly preferred embodiments, the host cells are modified so that they are deficient, or are naturally deficient, in an enzyme activity that is required for growth or survival of the cells in the presence of a selection agent and which is provided by the selectable marker. For example, Chinese Hamster Ovary (CHO) cells have been modified to be deficient for GS . In some preferred embodiments where vector includes a GS selectable marker, the host cell line is deficient in GS. In some particularly preferred embodiments, the GS deficient host cell line is the CHOZN® GS" / _cell line available from Merck KGaA. In other embodiments, where the selectable marker is, for example, DHFR, the cell line may preferably be deficient for DHFR activity (i.e., DHFR’). Suitable DHFR- cell lines include but are not limited to CHO-DG44 and derivatives thereof.The nucleic acid constructs and vectors of the present invention may be introduced into host cells by any suitable means such as by transfection, transformation or transduction. In some embodiments, after transfection or transduction, the cells are allowed to multiply, and are then trypsinized and re -plated. Individual colonies are then selected to provide clonally selected cell lines. In still further embodiments, the clonally selected cell lines are screened by Southern blotting or PCR assays to verify that the desired number of integration events has occurred. It is also contemplated that clonal selection allows the identification of superior protein producing cell lines. In other embodiments, the cells are not clonally selected following transfection.In some embodiments, the nucleic acid constructs encoding different proteins of interest are introduced into the host cells, for example by transfection or electroporation. The nucleic acid constructs encoding different proteins of interest can be introduced into the host cells at the same time or in a serial manner (e.g., a nucleic acid construct encoding a first protein of interest is introduced, a period of time is allowed to pass, and then a nucleic acid construct encoding a second protein of interest is introduced).In some embodiments of the present invention, following transformation of a suitable host strain and growth of the host strain to an appropriate cell density in media, the protein of interest is secreted during culture of the host cells. In some preferred embodiments where amplifiable markers are utilized, it is contemplated that culture of transduced host cells in amedium comprising an inhibitor of the gene. Suitable inhibitors include, but are not limited to methotrexate for inhibition of DHFR and methionine sulphoximinc (Msx) or phosphinothricin for inhibition of GS. It is contemplated that as concentrations of these inhibitors arc increased in a cell culture system, cells with higher copy numbers of the amplifiable marker (and thus the genes or genes of interest) or which contain higher-producing insertions are selected.Accordingly, the host cells containing vectors as described above are preferably cultured according to methods known in the art. Suitable culture conditions for mammalian cells are well known in the ail (See e.g., J. Immunol. Methods (1983) 56:221-234

[1983] , Animal Cell Culture: A Practical Approach 2nd Ed., Rickwood, D. and Hames, B. D., eds. Oxford University Press, New York

[1992] ).The host cell cultures of the present invention arc prepared in a media suitable for the particular cell being cultured. Commercially available media such as ActiPro media (HyClone), ExCell Advanced Fed Batch Medium (SAFC), Ham's F10 (Sigma, St. Louis, MO), Minimal Essential Medium (MEM, Sigma), RPMI-1640 (Sigma), and Dulbecco's Modified Eagle's Medium (DMEM, Sigma) are exemplary nutrient solutions. Suitable media are also described in U.S. Pat. Nos. 4,767,704; 4,657,866; 4,927,762; 5,122,469; 4,560,655; and WO 90 / 03430 and WO 87 / 00195; the disclosures of which are herein incorporated by reference. Any of these media may be supplemented as necessary with serum, hormones and / or other growth factors (such as insulin, transferrin, or epidermal growth factor), salts (such as sodium chloride, calcium, magnesium, and phosphate), buffers (such as HEPES), nucleosides (such as adenosine and thymidine), antibiotics (such as gentamycin (gentamicin), trace elements (defined as inorganic compounds usually present at final concentrations in the micromolar range) lipids (such as linoleic or other fatty acids) and their suitable carriers, and glucose or an equivalent energy source. In some preferred embodiments where selectable markers such as GS are utilized, for example, the media will lack glutamine. Any other necessary supplements may also be included at appropriate concentrations that would be known to those skilled in the art.The present invention also contemplates the use of a variety of culture systems (e.g., petri dishes, 96 well plates, roller bottles, and bioreactors) for the transfected host cells. For example, the transfected host cells can be cultured in a perfusion system. Perfusion culture refers to providing a continuous flow of culture medium through a culture maintained at high cell density. The cells are suspended and do not require a solid support to grow on. Generally, fresh nutrientsmust be supplied continuously with concomitant removal of toxic metabolites and, ideally, selective removal of dead cells. Filtering, entrapment and micro-capsulation methods arc all suitable for refreshing the culture environment at sufficient rates.As another example, in some embodiments a fed batch culture procedure can be employed. In the preferred fed batch culture the mammalian host, cells and culture medium are supplied to a culturing vessel initially and additional culture nutrients are fed, continuously or in discrete increments, to the culture during culturing, with or without periodic cell and / or product harvest before termination of culture. The fed batch culture can include, for example, a semi- continuous fed batch culture, wherein periodically whole culture (including cells and medium) is removed and replaced by fresh medium. Fed batch culture is distinguished from simple batch culture in which all components for cell culturing (including the cells and all culture nutrients) are supplied to the culturing vessel at the start of the culturing process. Fed batch culture can be further distinguished from perfusion culturing insofar as the supernatant is not removed from the culturing vessel during the process (in perfusion culturing, the cells are restrained in the culture by, e.g., filtration, encapsulation, anchoring to microcarriers etc. and the culture medium is continuously or intermittently introduced and removed from the culturing vessel). In some particularly preferred embodiments, the batch cultures are performed in roller bottles.Further, the cells of the culture may be propagated according to any scheme or routine that may be suitable for the particular host cell and the particular production plan contemplated. Therefore, the present invention contemplates a single step or multiple step culture procedure. In a single step culture, the host cells are inoculated into a culture environment and the processes of the instant invention are employed during a single production phase of the cell culture. Alternatively, a multi-stage culture is envisioned. In the multi-stage culture cells may be cultivated in a number of steps or phases. For instance, cells may be grown in a first step or growth phase culture wherein cells, possibly removed from storage, are inoculated into a medium suitable for promoting growth and high viability. The cells may be maintained in the growth phase for a suitable period of time by the addition of fresh medium to the host cell culture.Fed batch or continuous cell culture conditions are devised to enhance growth of the mammalian cells in the growth phase of the cell culture. In the growth phase cells are grown under conditions and for a period of time that is maximized for growth. Culture conditions, suchas temperature, pH, dissolved oxygen (dCb) and the like, are those used with the particular host and will be apparent to the ordinarily skilled artisan. Generally, the pH is adjusted to a level between about 6.5 and 7.5 using either an acid (e.g., CO2) or a base (e.g., Na^CCb or NaOH). A suitable temperature range for culturing mammalian cells such as CHO cells is between about 30° to 38° C and a suitable dO is between 5-90% of air saturation.Following the polypeptide production phase, the polypeptide of interest is recovered from the culture medium using techniques that are well established in the art. The protein of interest preferably is recovered from the culture medium as a secreted polypeptide (e.g., the secretion of the protein of interest is directed by a signal peptide sequence), although it also may be recovered from host cell lysates. As a first step, the culture medium or lysate is centrifuged to remove particulate cell debris. The polypeptide thereafter is purified from contaminant soluble proteins and polypeptides, with the following procedures being exemplary of suitable purification procedures: by fractionation on immunoaffinity or ion-exchange columns; ethanol precipitation; reverse phase HPLC; chromatography on silica or on a cation-exchange resin such as DEAE; chromatofocusing; SDS-PAGE; ammonium sulfate precipitation; gel filtration using, for example, Sephadex G-75; and protein A Sepharose columns to remove contaminants such as IgG. A protease inhibitor such as phenyl methyl sulfonyl fluoride (PMSF) also may be useful to inhibit proteolytic degradation during purification. Additionally, the protein of interest can be fused in frame to a marker sequence that allows for purification of the protein of interest. Nonlimiting examples of marker sequences include a hexa-histidine tag, which may be supplied by a vector, preferably a pQE-9 vector, and a hemagglutinin (HA) tag. The HA tag corresponds to an epitope derived from the influenza hemagglutinin protein (See e.g., Wilson et al., Cell, 37:767

[1984] ). One skilled in the art will appreciate that purification methods suitable for the polypeptide of interest may require modification to account for changes in the character of the polypeptide upon expression in recombinant cell culture.In some preferred embodiments, the nucleic acid constructs are incorporated into systems. In some embodiments, the systems comprise multiple nucleic acid constructs or vectors as described above which are intended for introduction into a host cell. In other preferred embodiments, the systems comprise one or more multiple nucleic acid constructs or vectors as described above which are intended for introduction into a host cell in addition to a nucleic acid or vector that encodes an enzyme that is necessary for incorporation of the nucleic acidconstructs into a host cell genome. Exemplary enzymes include, but are not limited to, transposes for use with transposon vector systems, integrases for use in systems which utilize integration sequences such as the PhiC31 system, MMLV systems, and the like, recombinases for use in vector systems such as Cre-loc, FLP-FRT and the like, and Cas9 nucleases for use in CRISPR based systems.EXPERIMENTALThe invention provides a unique way of combining the expression of Human DNase 1 and another product in a cell production system to allow for removal of DNA from the final product that is to be produced. The level of production of DNase may need to be varied depending on the product and its potential use. Human DNase 1 could be substituted for any other DNase or many other endonuclease molecules.Experiment 1: Production of CHO Cells Producing Human DNase 1The full-length Human DNase 1 coding DNA sequence was identified from NCBI GenBank and the following publication (Proc. Natl. Acad. Sci. USA, Vol. 87, pp. 9188-9192, December 1990), including the endogenous signal peptide region. The Human DNase 1 gene was designed, the flanking cloning sequences adding an optimized Kozak translation initiation sequence and added Hind III cloning site at the 5’ terminus and Xho I cloning site at the 3’ terminus of the human DNase 1 gene sequence for expedited expression cloning. See FIG. 1A for nucleic acid sequence (SEQ ID NO:1) and FIG. IB for protein sequence (SEQ ID NO:2). This sequence was also codon optimized for improved expression. This full-length CDS encoding human DNase 1 with the added flanking sequences was synthesized and cloned into pUC57 vector.The synthesized plasmid was digested with Hind III and Xho I restriction enzymes to release the Human DNase 1 encoding DNA fragment that was gel-purified and ligated into the pCS-newMCS-WPREplasmid digested with the same two enzymes. See FIGs. 2 and 3 for plasmid maps. The final clone was sequenced through the Human DNase 1 CDS to verify the congruity to the predicted DNA sequence, named pCS-CFSDl-WPRE (new ori).Table 1. Features of Starting Retrovector pCS-newMCS-WPRE (new ori)Table 2. Features of pCS-CFSDl-WPRE (new ori)1.1 Summary of development of DNase 1 expressing clones. Chinese Hamster Ovary (CHO-S) production cell lines were made by three rounds of transduction of the CHO-S parental cell line with retrovector made from the Human DNase 1 expression gene construct. The pooled populations were expanded for cryopreservation after each transduction.The (3X) pooled population was diluted into 96- well cell culture plates to establish clonal cell lines that originated from single cells. The clonal cell lines were screened by Pico Green DNA assay for protein titer. The top thirteen clones were expanded and tested in triplicate T175 flasks for overgrowth productivity and were cryopreserved. The QC analyses of the clonal cell lines were performed after cryopreservation. The QC tests included viability, gene copy index, retrovector component, bioburden, and mycoplasma. 1.2 Materials.The plasmid for expression of Vesicular Stomatitis Virus (Indiana) envelope glycoprotein (pHCMV-G) was originally developed by Pangcnix (San Diego, CA) and was prepared by Bayou Biolabs (Harahan, LA). The retrovector plasmid was prepared in house by endotoxin free maxi-prep (Qiagen, Valencia, CA) using the manufacturer’s protocol. Quality control analyses included determination of DNA concentration and sequencing of the protein coding region and cloning junctions. DNA sequencing was performed by ACGT, Inc. (Wheeling, IL) using primers manufactured by Invitrogen (Carlsbad, CA).The retrovector packaging cell line 293GP was developed by Pangenix (San Diego, CA), and has been characterized and master cell banked at Bio Reliance (Rockville, MD) for CPS-M. The suspension-adapted Chinese Hamster Ovary cell line (CHO-S) was received from GIBCO Life Technologies Inc. (Rockville, MD, Catalog #11619012), and has been characterized and master cell banked at BioReliance (Rockville, MD) for CPS-M. DF medium is Dulbecco’s Modified Eagle’s Medium (HyClone, Catalog # SH30243) plus 10% Fetal Bovine Serum (FBS) from HyClone (Catalog # SH30070). DFP medium is DF medium plus 10 pg / mL phleomycin. PF CHO Liquid Soy Medium (PF CHO LS) is purchased from HyClone (Catalog # SH30359). 1.3 Methods.1.3.1 Retrovector production.293GP cells were cultivated in DFP medium and then passaged to 16 T150 flasks using trypsin (HyClone Catalog # SH30042). Two hours prior to transfection, the flasks were changed to 25 mL of DF medium. Transfection was performed following SOP STM-CEL-0325 using 864 pg of retrovector construct DNA plasmid pCS-CFSDl-WPRE (new ori), and 54 pg of expression plasmid for Vesicular Stomatitis Virus envelope glycoprotein. The plasmid solutions were combined with 1:10 TE for a total volume of 17.47 mL and 2.52 mL of 2M CaCh, and then precipitated by dropwise addition into 19.92 mL of 2X HBS solution. Then, 2.5 mL of suspension was added to each of the 16 T150 flasks and incubated on the cells for six hours at 37°C in a 5% CO2 atmosphere. Growth at 37°C ± 1°C in a 5% ± 1% CO2 atmosphere will be referred to as standard conditions from this point forward.After six hours, the culture medium was replaced with 20 mL of fresh DF medium. The flasks were incubated under standard conditions until the second day after transfection. The medium was collected from the 16 T150 flasks and filtered through a 0.45 micron filter and then a 0.2 micron filter. The retrovector was concentrated from the 320 mL of harvested medium bycentrifugation in a Beckman J-30I centrifuge with a JA-30.5 rotor at 18,750 rpm (40,000 x G) for 90 minutes at 4°C. The supernatant was aspirated from the centrifuge tubes and the pelleted material in each tube was resuspended in 25 pL of PF CHO LS medium. The concentrated vector was used for the CHO-S transduction step.1.3.2 Transduction of CHO-S cells with retrovectorsParental CHO-S cells were established in culture and prepared for transduction following procedure SOP STM-CEL-0350. A suspension of 4 x 104viable CHO-S cells was prepared in 5 mL of PF CHO LS medium with 8 pg / mL polybrene. This cell suspension was incubated under standard conditions for a minimum of two hours prior to the addition of the retrovector. Immediately prior to the addition of retrovector, the cell suspension was centrifuged for four minutes at 1500 rpm (500 x G) in a tabletop centrifuge (Beckman Coulter Allegra 6 with a GH 3.8 rotor) and the supernatant was removed without disturbing the cell pellet. The retrovector was added to the CHO-S cell pellet, mixed and was incubated under standard conditions.After one day, 5 mL of PF CHO LS medium was added to the tube containing the cell- retrovector mixture and mixed. This mixture was then centrifuged for four minutes at 1500 rpm (500 x G) in a tabletop centrifuge. The supernatant containing any residual retrovector was removed and another wash and spin were performed. The supernatant was removed without disturbing the cell pellet, and the cells were resuspended in 2 mL PF CHO LS medium. The cell suspension was transferred to a 12- well cell culture plate and expanded through consecutive passages into successively larger cell culture flasks using PF CHO LS medium.Each subsequent transduction was performed following the same methods as before using cells in culture from the previous transduction. The pooled population from each transduction was expanded and cryopreserved. In addition, a sample of cells was submitted for gene copy index testing.1.3.3 Establishment of clonal cell linesClonal selection was performed on an aliquot of sCHO-S / sC-CFSDl-R (3X) cells at passage level 6. The cells were diluted to 0.5 and 0.75 viable cells per 200 pL in PF CHO LS medium with 2% FBS. The addition of FBS allowed cells to grow as adherent cultures and assisted in the growth of founder colonies originating from single cells. Twenty 96-well plates were seeded with 200 p L per well of cell suspension for each of the dilutions.1.3.4 Selection and testing of top clonal cell lines.The seeded 96-well plates were incubated under standard conditions and were observed microscopically on two different days for the development of colonics originating from single parental cells (SOP STM-CEL-0330). Media was collected on Day 14 from 218 wells in which single colonies were observed. The media samples were screened by an activity assay to determine protein titer. Media was replaced with PF CHO LS without FBS and cells were cultured in PF CHO LS without FBS from this point on. The top 24 clones were selected based on Human DNase 1 titer. Eleven clones did not survive the transition from adherent to suspension and were discarded. The remaining 13 clones were expanded for productivity testing in an overgrowth study. Triplicate T 175 flasks were seeded with 300,000 viable cells per mL in 50 mL working volume of PF CHO LS. Viable cell densities (VCD) were determined on day three and samples were collected for protein analysis by an activity assay on days three and 14. The cultures were terminated on day 14.1.4 Results and discussion.The Human DNase 1 expression cell lines were created by performing multiple rounds of transduction of the sCHO-S parental cell line using retrovector made with construct DNA plasmid for the expression of pCS-CFSDl-WPRE (new ori). The pooled population was expanded for cryopreservation after each transduction and a sample of cells was submitted for gene copy analysis. The names used to identify the sCHO-S cell lines after each round of transduction and the gene copy index results are shown.Table 3. Gene Copy Index Results of the Human DNase 1 Expression Cell LinesEach transduction created a population of cells with the Human DNase 1 genes inserted in differing numbers and locations in the production cell line. Clonal selection by limited dilution plating was performed to identify candidate clonal cell lines for production. The pooled population of sCHO-S / sC-CFSDl-R (3X) cells was diluted to 0.5 and 0.75 viable cells per 200 pL media and plated in 96-well cell culture plates to establish clonal cell lines that originatedfrom single cells. Forty 96-well plates were screened twice microscopically to identify 218 wells with a single colony.In the clonal selection procedure, the number of wells containing single cells corresponds with the expected number based on seeding density; however, the number of wells with two or more cell colonies is usually higher than the number of wells with single cells. The screening process is very stringent and as such, any well that contains a colony slightly outside the ‘norm’ is classified as a 2+ cell-colony well (i.e. a larger than normal colony, satellite cells, irregular shaped colony, floating cells). Due to this selection process, a number of what are probably single cell colonies are categorized as > 1 cell colonies per well.Table 4. Microscopic Clonal Screening ResultsFourteen days post seeding, media samples were collected from the 96-well plates and screened by an activity assay for protein production. The top 24 clones from the 96-well plate samples ranged from 32 to 47 g / mL of Human DNase 1 protein.Table 5. Activity Assay Results from 96-Well Plates*Discarded due to poor growth.The top 13 clones were set up in triplicate T175 flasks for overgrowth productivity testing.Table 6. Productivity Results of the Top 12 Clones in Triplicate T175 Flasks1.5 Conclusion.Clonal cell lines were made by performing limited dilution cloning of the sCHO- S / sC-CFSDl-R (3X) pooled population. Twelve clonal cell lines were screened by overgrowth analysis. Protein titers were determined by ELISA. The average maximum protein level of the top 12 clones was 40 mg / L. The top five clones, #1, 20, 66, 85 and 98 were primarily selected based on titer. These top five clonal lines had maximum expression levels ranging from 42 mg / L for clone #85 to 47 mg / L for clone #1. The mean PCD of the top 12 clones was 8.43 on day three. The PCD for the top five clones ranged from 10.64 for clone #66 to 16.67 for clone #98. The clones produced high levels of Human DNase 1 and the production of the product showed no effects on CHO cell growth or behavior. Upon further cell culture optimization levels of protein production reach as high as 2 g / L in fed- batch culture.Experiment 2: Production of CHO cells expressing Human DNase 1 and an antibody productThe presence of residual DNA during the culture of cells, may in some instances inhibit the production of particular recombinant proteins or in other instances bind so tightly to the recombinant protein product that prevents effective removal of the DNA during purification of the product. The co-expression of Human DNase 1 or other nucleases with these types of products, may improve product expression and product purity. In this experiment we coexpressed Human DNase 1 at different levels with an antibody product to determine if this approach could be successful. The GPEx Lightning technology was utilized to co-express Human DNase 1, an antibody heavy chain and an antibody light chain in CHO cells.2.1 Summary.Based on the obtained antibody sequence information, Catalent designed coding DNA sequences (CDS) for optimal expression of the antibody light chain (LC) and heavy chain (HC) of GMAB, as well as the Human DNase 1 gene. Each of the three genes were then cloned into the GPEx® Lightning expression vector. The LC and HC CDS in the expression vector (plasmid) were each confirmed by DNA sequencing.Nine independent production cell lines were made by performing a single round of transfections of the 1F7 CHO parental cell line with the gene constructs developed to express GMAB and Human DNase 1. The nine pooled populations underwent selection via removal of glutamine and were expanded for gene copy index value (GCIV) testing and cryopreservation. The CHOZn / GA04-LC / HC / DNase pooled population #6 cell line exhibited the most desirable ratio of GCI values between LC, HC, and DNase 1, and was further tested in a fed-batch productivity study.The CHOZn / GA04-LC / HC / DNase pooled population #6 was processed through a single round of clonal selection using the Beacon® platform to establish clonal cell lines that originated from a single cell expressing the protein of interest. The clones reported herein have an average calculated probability of monoclonality of > 99%. The Top 12 clones were grown in a fed-batch productivity culture study under generic conditions to delineate the highest GMAB-producing cell lines.Clones #31, 37, and 1328 were identified as the top three Master Cell Bank (MCB) candidates, with Clone #11 as a backup candidate, based on protein production and cell line quality.2.2 DNA constructions and cloning.2.2.1 Materials and methods.Based on starting antibody sequence information, we designed CDS for optimal expression of the antibody LC and HC. The unique CDS for the GMAB LC and HC were designed by Catalent using proprietary Triplet-Fix® codon optimization technology and public access sources including NCBI and IMGT websites. Unique restriction sites for cloning the complete LC and HC CDS into Catalent expression vectors were added on the 5’ and 3’ ends of the CDS. Additionally, a Kozak sequence for efficient protein expression and two tandem stop codons to prevent translational read-throughs were introduced at the 5’ and 3’ ends, respectively. Human DNase 1 “Pathway” was to be co-expressed with the antibody chains to potentially enhance their expression and secretion. The Human DNase 1 CDS was subcloned into Lightning expression vector as described.Each designed CDS was synthesized by IDT. For each full-length CDS, cloning was performed in a similar manner. The IDT gBlock® or the parental vector was digested with Hindlll and Xhol (HC and Pathway) or Notl and Bglll (LC) restriction endonucleases and the released CDS was ligated into the GPEx® Lightning expression vector that had also been digested with the same two corresponding restriction endonucleases. The ligation reactions were performed according to the manufacturer’s procedure using NEBuilder HiFi DNA Assembly Master Mix. The Lightning expression vector 207pucl9attB287-GS-NewMCS- WPRE-TKpa was previously generated at Catalent.2.2.2 Results.A clone was confirmed to encode full-length GMAB LC and HC CDS and the DNase Pathway CDS. The new plasmid encoding each of the three CDS were named and designated respectively: 207attB-GS-h3E10LC-WPRE and, 207attB-GS-h3E10HC-WPRE, and 215- pucl9attB287-GS-Pathway-WPRE-TKpa. All three CDS and the flanking DNA cloning junctions in the final expression constructs are shown. The cloning restriction sites are also shown. The expression vectors were DNA-sequenced through all three CDS and the cloning junctions to exclude mutations.FIG. 4 provides the plasmid map for plasmid 207attB-GS- h3E10LC -WPRE. FIG. 5 provides the plasmid map for plasmid 207attB-GS-h3E10HC-WPRE. FIG. 6A provides the nucleic acid sequence for the 1008-215 -Pathway-207 insert (SEQ ID NO:3) while FIG. 6B provides the amino acid sequence for the 1008-215-Pathway-207 insert (SEQ ID NO:4). FIG. 7 provides the plasmid map for plasmid 215-pucl9attB287-GS-Pathway-WPRE-TKpa. Tables 7, 8 and 9 provide a summary of the features of these expression vectors.Table 7. Features of Expression Vector 207attB-GS-h3E10LC-WPRETable 8. Features of Expression Vector 207attB-GS-h3E10HC-WPRETable 9. Features of Expression Vector 215-pucl9attB287-GS-Pathway-WPRE-TKpa2.3 Cell line development2.3.1 Materials and methods.Next, parental GPEx Lightning 1F7 CHO cells were established in culture and prepared for transfection. EX-CELL® Advanced CHO Fed-Batch medium supplemented with6 mM L-glutamine was used as the base media for culture maintenance. Immediately prior to the addition of transfection reagents and DNA, a required amount of 1F7 parental cell suspension was centrifuged for five minutes at 500 x g in a tabletop centrifuge (Sorvall Legend XT with TX-750 rotor). As much supernatant as possible was removed without disturbing the cell pellet. Cells were then washed with 5 mL of CHOGro expression media, spun at 500 x g in a tabletop centrifuge, and aspirated a second time. Fresh CHOGro expression media was then added to the cell pellet to achieve a final suspension of 200 x 105viable 1F7 cells in 2 mL of CHOGro expression medium. The LC / HC / DNase 207pucl9attB287-GS-NewMCS-WPRE-TKpa plasmids were combined at ratios of 1:1 :1, 5:5:1, and 10:10: 1 (LC:HC:DNase) with the total amount of DNA remaining constant across the 3 DNA preparations. DNA plasmids and the recombinase DNA plasmid were mixed in Optipro media at a ratio of 500:1 in a tube (tube #1). The expifectamine reagent was diluted in Optipro media in a second tube (tube #2). Tubes #1 and 2 were mixed, allowed to sit for 1- 5 min, and then added to the 1F7 cells (30 x 105) in a 50 mL culture vessel and then shaken at 250 rpm and 37 °C. Three independent transfections were performed resulting in three individual pooled populations.After 2-4 hours of incubation, the cell-transfection reagent mixture was centrifuged for five minutes at 500 x g in a tabletop centrifuge. The supernatant was removed, and the transfected cells were suspended in 10 mL of EX-CELL® Advanced CHO Fed-Batch medium supplemented with 2% ACF and 6 mM L-glutamine. The transfected cells recovered for three days post-transfection before the selection process was initiated. After the cells had fully recovered from the transfection process, cultures were centrifuged for five minutes at 500 x g in a tabletop centrifuge; all media was aspirated without disturbing the pellet, and cells were resuspended in 10 mL of EX-CELL® Advanced CHO Fed-Batch medium without L- Glutamine or ACF. The cultures were then counted at least every other day to monitor the viable cell density, the percent viability, and the doubling time. Generally, the transfected cells exhibited a decline in viability from the beginning of selection until Day 10 posttransfection (Nadir). After the minimum viability was achieved, the selection process was complete, and successfully transfected cells began to recover. Cultures recovered for another 10 days until the viability fully recovered and the doubling time stabilized.The pooled cell populations from transfection were expanded and cryopreserved. A supernatant sample was taken for protein titer analysis by BioHT and protein quality analysis by SDS-PAGE. A sample of the cells was submitted for gene copy index (GCIV) analysis and residual recombinase analysis.2.3.2 Generation of GMAB expression cell line.Next, the GMAB expression cell lines were created by performing one round of transfection of the 1F7 parental cell line with the 3 DNA plasmids. The names used to identify each of the nine 1F7 cell lines after transfection, along with the GCIV results are shown. See Table 10. The SDS-PAGE results are also shown. See Fig. 8.Table 10. Transfection of the GMAB Expression Cell Lines2.4 Fed-batch production from a pooled cell population.2.4.1 Materials and methods.Cell line CHOZn / GA04-LC / HC / DNase pooled population #6 was scaled up for initial protein production by fed-batch analysis in 500mL Shake Flask seeded at 6.0 x 105viable cells / mL in 120 mL of ActiPro medium supplemented with 4% ps307.Viable cell density (VCD) and viability was determined every day after Day 2. Protein titer was determined by BioHT IgG analysis. Specific protein production (picograms produced per cell per day, ped) was determined by taking the slope of the linear fit curve of protein concentration (in pg / nL) plotted against integral cell density (cells* day / nL). The culture was terminated when the viability fell below 90%, but did not exceed Day 14. See FIGs. 9-11.2.4.2. Results.The gels showed that each of the three protein products (LC / HC / DNase 1) were expressed in the expected ratio based on the plasmid transfection. The production of DNase 1 by the cells does not appear to inhibit the production of functional antibody at high levels. These cells behaved similarly to a standard CHO cell line just producing an antibody. The CHOZn / GA04-LC / HC / DNase pooled population reached a peak viable cell density on Day 10 of 254 x 105cells / mL, and was harvested on Day 14 with a viability of 96.4%. The cell line produced 5.1 picograms of antibody / cell / day (ped).2.5 Clonal selection with the Beacon Instrument.2.5.1 Materials and methods.One round of clonal selection was performed using the Beacon instrument (Berkeley Lights Inc (BLI), Emeryville, CA) to identify candidate clonal cell line. The Beacon optofluidic platform uses OptoSelect™ Opto-Electric Positioning technology (light cages) to place single cells into 1 nL pens on chips. The instrument pens, cultures, assays, images, ranks, and then exports high-expressing clonal cell lines. CHOZn / GA04-LC / HC / DNase cells were seeded at 1-2 million cells per mL in a volume of 10 mL in dynamic culture, 1 day prior to loading on the Beacon instrument that contained G12.1 medium supplemented with 6 mM L-glutamine and 0.25 g / L PS307 with 2.5% animal component free (ACF) supplement. The viable cell density of the cell suspension upon loading was 38.2 x 105cells / ml and the viability was 99.9%. The penning efficiency (percentage of pens containing single cells) was calculated to be 80.7% (2837 / 3516). Each pen was verified manually by an operator to ensure a single cell was present. Cells were then cultured on the chip for three days in G12.1 medium supplemented with 6 mM L-glutamine and 0.25 g / L PS307 with 2.5% ACF supplement. It was confirmed that no pens were overgrown (not more than 50% full). Thepercent on chip clonal expansion (OCCE; percentage of pens containing 4 or more cells by Day 3) was calculated to be 96.3% (2731 / 2837).On Day 3 of the culture, an in-pen diffusion assay using Spotlight Hu3™ (BLI) was performed, and parameters were set and applied for the selection of colonies to be exported. Clones fitting the criteria were ranked by titer score. The Top 96 colonies were exported into optical 96-well plates (VWR, Cat. #82050-772) with 160 pL per well of G12.1 culture medium with 6 mM L-glutamine and 0.25 g / L PS307 with 2.5% ACF supplement.Colonies were cultured for eleven days and imaged via the Operetta Imaging system (Perkin Elmer) at Days 3, 7, 9, and 10 post-export for the purposes of monitoring growth. Clones were expanded from 96-well plates into 24-well plates (VWR, Cat. #82050-846) and then into 6-well plates. Clones were submitted for gene copy index value (GCIV) analysis. Subsequently, the clones were expanded into shaking culture in 50mL bioreactor tubes, and then up to a volume of 120 mL in 500mL shake flasks for cryopreservation. Cell counts were performed at each passage of dynamic culture. Twenty-three vials of each cell line were prepared for cryopreservation; three vials of each clone were used for QC testing. The QC tests included viability upon thaw, MET (bioburden), and mycoplasma.Table 11. Beacon® Parameters Used for Selection of Clones for Export2.5.2 Clonal analysisThe transfection process creates a population of cells with the GMAB CDS inserted in variable quantities and locations in the production cell line. During clonal selection, a total of approximately 2731 colonies were evaluated.The Top 96 clones ranked by AU score were exported into optical well 96-well plates. These clones were observed for growth using the Operetta imaging system (Perkin Elmer) on Days 3, 7, 9, and 10 post-export. Of the Top 96 clones, a total of 74 scaled up successfully from 96-well plates, to 24-well plates, and then to 6-well plates. The Top 48 clones selected by GCI values were next evaluated in a fed-batch productivity study.Table 12. Beacon Results - Top 71 Clones2.5.3 Productivity study in 24-well plateThe Top 71 clones as ranked by GCI values were tested in a dynamic batch culture in 24-well Deep Well Plates and evaluated for VCD and protein titer on Days 3 and 6 to identify the Top 12 clones for cryopreservation and further characterization in an ambrl5 study. The VCD, viability, and titer for the 48 clones were determined. See Table 13 and FIGs. 12-14.Table 13. 24-well Plate Specific Production Rates and Harvest Data2.5.4 Testing and Selection of the Clonal Cell Lines in an Ambrl5™.The selected Top 12 clonal cell lines were tested for productivity in an Ambrl5™ miniature bioreactor system. The Ambrl5™ mimics the operational parameters of classical bioreactors (2 L and 10 L) at a smaller scale (10-15 mF). The automated workstation is comprised of blocks of 12 disposable microbioreactors with independent environmental (aeration and pH), feed, and sampling controls for each culture. The clones were evaluated in ActiPro medium with Cell Boost™ 7a / 7b feed and ExCell Advance Fed-Batch medium with Feedl / 4FEED, with a temperature shift to 34 °C occurring on Day 6 for both the ActiPro and Fed-Batch media cultures. Cultures were terminated when the viability fell below 90% and did not exceed Day 14.For scale-up, cells were used from the CHOZn / GA04-LC / HC / DNase cell line for Clones #5, 11, 12, 28, 31, 37, 66, 317, 419, 1328, 1476, and 1730. The cells were maintained in Actipro medium supplemented with 0.25 g / L PS307; cells from each of the clonal cell lines were then adapted to Fed-Batch medium (with no supplements). Three passages were performed on the Top 12 clones in ActiPro medium or Fed-Batch medium prior to seeding in the Ambrl5™ vessels.The clones were inoculated in Ambrl5™ vessels at cell densities of 6.0 x 105cells / mL or 10.0 x 105cells / mL. Samples were taken daily for VCD, viability, and metabolite levels (glucose, lactate, and ammonia). Titer was measured on days 4, 8, 12, 14, or harvest.Table 14. AmbrlS™ Experimental DesignThe results comparing the clone number relative to the cumulative cell density (CCD) are shown, the clone number relative to the titer i, and the clone number relative to rQp.Based on the analysis of the Ambrl5™ production data, the Top 4 clonal candidates are Clones #31, 37, 1328, and 11 based on titer production (BioHT).On average, Fed-Batch conditions yielded higher titers than Actipro conditions, however the highest overall titer was observed in ActiPro. Further optimization of these conditions and other culture conditions could result in additional protein production.2.6 Overall conclusionsThe results indicate that co-expression of a nuclease in this example Human DNase 1 and a second recombinant protein or antibody is possible. The production for the antibody in these cell lines was at commercially viable levels. Based on an activity assay being performed on media harvested from the culture, the Human DNase 1 is active and capable of cleaving DNA in the environment of cell culture media. We have shown the level of DNase 1 being produced can be adjusted in order to obtain the desired effect for that particular product.Experiment 3: A Comparison of Stable CHO Cell Pools Expressing the 3E10 Antibody with and without Human DNase 1

[0001] The GPEx Lightning Process was used to generate two stable cell pools expressing the 3E10 antibody. One cell pool, PP5, was designed only to express the heavy and light chain of the 3E10 antibody. The second cell pool, PP6, was designed to express the heavy and light chain of the 3E10 antibody and the human DNase 1 enzyme. After generating the two stable cell pools, they were tested for heavy and light chain gene copy index values and for productivity performance in fed-batch culture.Table 15. Heavy and Light Chain Gene Copy Index for PP5 and PP6

[0002] The results indicated that the gene copy index values are similar for both the PP5 and PP6 cell pools. Fed-batch productivity studies were performed for each of the cell pools at Day 14. A comparison of the results for PP5 and PP6 is shown below:Table 16. Fed-batch Productivity Studies for PP5 and PP6 at Day 14

[0003] The results indicated that 3E10 antibody expression alone produced much lower- than-expected titers for an antibody product and the cells producing the antibody showed poorer growth characteristics when compared to cells producing a traditional antibody product. Since the 3E10 antibody binds to nucleic acids forming a complex that may be internalized by cells, it was hypothesized that DNA from dead cells in culture may be binding to secreted antibody and entering cells causing issues with expression and cell growth. It was further hypothesized that removal of DNA by having the cells secrete human DNase 1 in addition to the 3E10 antibody may improve antibody production and cell health. The comparison of the two cell pools, PP5 and PP6, support these hypotheses. The GPEx Lightning technology gene copy index is directly correlated to product titer. For the two stable pool cell lines, the PP5 (the pool without DNase 1) would be expected to produce moreantibody than PP6 (the pool with DNase 1) since it has a slightly higher transgene number for both heavy and light chain genes. The opposite result was observed, with the PP6 cell pool producing 3-fold higher antibody levels than PP5 cell pool. Also, the cell growth behavior of the PP6 cell pool is more like CHO cells producing other types of antibodies than the PP5 cell pool. These results indicate that the DNase 1 co-expression approach is a much more viable method to produce the 3E10 antibody than the traditional method of just heavy and light chain expression in CHO cells.Experiment 4. Expression of antibody heavy and light chains at different ratios.This example provides data examining the use of the GPEx Lightning process for producing an antibody product “Yourway” using two methods. The first method with both the Yourway heavy and light chain genes on the same expression plasmid and the second with the heavy and light chain genes on different plasmids. We also examined how varying the ratios of those heavy and light chain plasmids during the GPEx Lightning process would affect antibody expression, protein quality and cell growth characteristics. For each of GPEx Lightning transfections 2 micrograms of plasmid was used. In the case of the different ratios of heavy to light chain plasmid, the amounts were 1) 1.8 micrograms heavy chain - 0.2 micrograms of light chain; 2) 1.6 micrograms heavy chain - 0.4 micrograms of light chain; 3) 1.4 micrograms heavy chain - 0.6 micrograms of light chain; 4) 1.2 micrograms heavy chain - 0.8 micrograms of light chain; 5) 1.0 micrograms heavy chain - 1.0 micrograms of light chain; 6) 0.8 micrograms heavy chain - 1.2 micrograms of light chain; 7) 0.6 micrograms heavy chain - 1.4 micrograms of light chain; 8) 0.4 micrograms heavy chain - 1.6 micrograms of light chain; 9) 0.2 micrograms heavy chain - 1.8 micrograms of light chain. In general, after the GPEx Lightning process, the number of each of the genes inserted into the pooled cell line followed the amount of plasmid associated with the transfection proportionally. The subsequent protein expression of each chain followed a similar trend. However, since most antibody heavy chains can be toxic to cells when a light chain is not present to bind to it during production, problems with cell growth were seen at the high ratios of heavy chain. That fact, and the GPEx Lightning process that uses a unique GS selection process appears to have skewed the high heavy chain ratio pools slightly towards the cells in that pool producing more light chain, even in the cell lines with a high ratio of heavy chain. The highest producing ratios were those that were producing extra light chain as observed in the non-reducing SDS-PAGE gels and were around the 1:1 ratio of the two chains. Sincelight chain genes are smaller, in general more light chain mRNA is produced / gene inserted and more light chain protein is translated per mRNA produced.Experimental Design and ResultsGene constructs described herein were used as part of the GPEx Lightning process for development of the antibody heavy chain and light chain ratio pooled cell lines. The cell lines were produced using these gene constructs and standard GPEx Lightning methodology.The expression sequence for the Yourway antibody light chain using the bovine alpha-lactalbumin signal peptide for secretion from the cell is provided in FIG. 15. The Yourway antibody light coding DNA sequence (CDS) and protein sequence are shown. The signal peptide included into the CDS is set off in a dark blue color.The Yourway light chain expression vector used for the GPEx Lightning transfection process is provided in FIG. 16.The expression sequence for the Yourway antibody using the bovine alphalactalbumin signal peptide for secretion from the cell is provided in FIG. 17. The Yourway antibody heavy coding DNA sequence (CDS) and protein sequence are shown. The signal peptide included into the CDS is set off in a dark blue color.The Yourway heavy chain expression vector used for the GPEx Lightning transfection process is depicted in FIG. 18.The Yourway heavy chain and light chain (two chain) expression vector used for the GPEx Lightning transfection process is depicted in FIG. 19.The GPEx Lightning technology uses a recombinase to specifically insert genes into “dock” sequences recognized by the recombinase and placed into the cell line using the GPEx process. Approximately 150-200 “dock” sequences are present in the cell line and available for gene insertion. Nine different cell pools were produced with the GPEx Lightning process using the gene constructs shown in figures 1-5. The pooled cell lines contained the following plasmid ratios / amounts . In the case of the different ratios of heavy to light chain plasmid, the amounts were 1) 1.8 micrograms heavy chain - 0.2 micrograms of light chain; 2) 1.6 micrograms heavy chain - 0.4 micrograms of light chain; 3) 1.4 micrograms heavy chain - 0.6 micrograms of light chain; 4) 1.2 micrograms heavy chain - 0.8 micrograms of light chain; 5) 1.0 micrograms heavy chain - 1.0 micrograms of light chain; 6) 0.8 micrograms heavy chain - 1.2 micrograms of light chain; 7) 0.6 micrograms heavy chain - 1.4 micrograms of light chain; 8) 0.4 micrograms heavy chain - 1.6 micrograms of light chain; 9) 0.2 micrograms heavy chain - 1.8 micrograms of light chain. As part of the GPEx Lightningprocess after transfection of the integrase and the transgene constructs, selection by removal of glutamine from the media occurs to allow only cell lines containing the transgene (and the glutamine synthase) to survive. How fast cell lines recover from the selection can typically indicate how well the process worked as well as if there are any issues caused by transgene expression.The pooled cell lines that contained a higher ratio of heavy to light chain gene showed slower recoveries during the selection process. Highlighting the potential toxicity of the heavy chain when not paired with a light chain. As the ratios became more favorable, the cells recovered faster.Attorney Docket No. CATA-41307.601Data table for the different heavy chain to light chain ratios.The number of gene copies was measured using a value called gene copy index. This is not actual gene copy number but a ratio of each gene to an endogenous CHO control gene. The details of the assay are shown in the appendix. The gene copy index of the pools followed the expect trend of the ratio transfected into the cells. However it can be seen that at high ratios of heavy chain the numbers are slightly skewed away from extra heavy chain genes due to the potential toxicity and the GS selection that occurs with the process when you compare the values when light chain is in significant excess. Fed-batch productivity studies were performed on each of the cell pools in duplicate with the results averaged. The best antibody yields for this particular antibody were associated with transfection ratios of light chain to heavy chain of 1 : 1 or 1.2:0.8 where light chain is in slight excess. As was observed in the cell growth data, the higher heavy chain ratios grew to lower cell densities and had lower IVCD’s.FIG. 20 provides a graph showing cell viability curves after glutamine selection. Pooled cell line productivity data for each of the pool duplicates with the double chain (Light + Heavy chain) construct as the control (CT36-2) is provided in FIG. 21. Productivity of some of the ratio pools reached similar or higher levels as the control construct.SDS-PAGE gel analysis of the of the pools showed the expected trends is provided in FIG. 22. More product missing light chain in the pools with higher ratios of heavy chain and much more free light chain in the pools with higher light chain ratios. The control pool with both genes on the same gene construct, showed free light chain similar to the 1 : 1 ratio cell pools, as was expected since the number of each gene should be in a similar balance. This supports the efficiency at which the gene insertion occurs and how the ratio of genes inserting is directly related to the amount of each plasmid in the initial transfection reaction.All publications and patents mentioned in the above specification are herein incorporated by reference. Various modifications and variations of the described method and system of the invention will be apparent to those skilled in the art without departing from the scope and spirit of the invention. Although the invention has been described in connection with specific preferred embodiments, it should be understood that the invention as claimed should not be unduly limited to such specific embodiments. Indeed, various modifications of the described modes for carrying out the invention that are obvious to those skilled in the field of this invention are intended to be within the scope of the following claims.

Claims

CLAIMSWhat is claimed is:

1. A eukaryotic host cell comprising a first exogenous nucleic acid sequence encoding DNase 1 operably linked to a promoter sequence and at least a second exogenous nucleic acid sequence encoding a first product of interest operably linked to a promoter sequence, wherein the first sequence encoding DNasel and the at least second exogenous encoding the first product of interest are co-expressed.

2. The host cell of claim 1, wherein the host cell further comprises a third exogenous nucleic acid sequence encoding a second product of interest.

3. The host cell of any one of claims 1 to 2, wherein the host cell further comprises a fourth exogenous nucleic acid sequence encoding a third product of interest.

4. The host cell of any one of claims 1 to 3, wherein the host cell further comprises a fifth exogenous nucleic acid sequence encoding a fourth product of interest.

5. The host cell of any one of claims 1 to 4, wherein the host cell further comprises a sixth exogenous nucleic acid sequence encoding a fifth product of interest.

6. The host cell of any one of claims 1 to 5, wherein the one or more products of interest is a protein or proteins.

7. The host cell of any one of claims 1 to 6, wherein the sequence encoding DNasel is further operably linked to a secretion signal sequence.

8. The host cell of claim 1, wherein the first product of interest is selected from the group consisting of an immunoglobulin heavy chain sequence and an immunoglobulin light chain sequence.

9. The host cell of claim 1, wherein the host cell further comprises a third exogenous nucleic acid sequence encoding a second product of interest operably linked to a promotersequence and a secretion signal sequence, wherein the first product of interest is an immunoglobulin light chain sequence and the second product of interest is an immunoglobulin heavy chain sequence.

10. The host cell of any one of claims 1 to 5, wherein the one or more products of interest are a nucleic acid or nucleic acids.1 1 . The host cell of claim 10, wherein the at least a second exogenous nucleic acid is a viral nucleic acid.

12. The host cell of claim 11 , wherein the viral nucleic acids are selected from the group consisting of retroviral nucleic acids, lentiviral nucleic acids, and AAV nucleic acids.

13. The host cell of claim 10, wherein the at least a second exogenous nucleic acid is an exosomal nucleic acid.

14. The host cell of any one of claims 1 to 13, wherein the first exogenous nucleic acid sequence and at least a second exogenous nucleic acid sequence are stably integrated into the genome of the host cell.

15. The host cell of any one of claims 1 to 14, wherein from 2 to 500 copies each of the first exogenous nucleic acid sequence and at least a second exogenous nucleic acid sequence are stably integrated into the genome of the host cell.

16. The host cell of any one of claims 1 to 14, wherein from 5 to 200 copies each of the first exogenous nucleic acid sequence and at least a second exogenous nucleic acid sequence are stably integrated into the genome of the host cell.

17. The host cell of any one of claims 1 to 16, wherein the ratio of the first exogenous nucleic acid sequence to the second exogenous nucleic acid sequence is from 1: 1 to 1 :100.

18. The host cell of any one of claims 1 to 16, wherein the ratio of the first exogenous nucleic acid sequence to the second exogenous nucleic acid sequence is from 1:2 to 1 : 100.

19. The host cell of any one of claims 1 to 18, wherein the first exogenous nucleic acid sequence and at least a second exogenous nucleic acid sequence are stably integrated into the genome of the host cell at a docking site.

20. The host cell of claim 19, wherein the docking site comprises at least one dock site insertion element and the exogenous nucleic acid sequences each comprise at least one insertion element compatible with the at least one dock site insertion element in the integrated docking sites.

21. The host cell of any one of claims 19 to 20, wherein the exogenous nucleic acid sequences further comprise at least the following elements in operable association in 5’ to 3’ order: an internal promoter sequence; a nucleic acid sequence encoding the first product of interest or second product that is operably linked to the internal promoter; and a poly A signal sequence.

22. The host cell of claim 21, wherein the exogenous nucleic acid sequences comprise a selectable marker sequence.

23. The host cell of claim 22, wherein the exogenous nucleic acid sequences comprise different selectable marker sequences.

24. The host cell of claim 21 , wherein one of the exogenous nucleic acid sequences comprises a selectable marker sequence and the other of the exogenous nucleic acid sequences does not comprise a selectable marker sequence.

25. The host cell of any one of claims 22 to 24, wherein the selectable marker sequences are 5 ’ to the internal promoter sequence and are operably linked to a 5 ’ promoter sequence.

26. The host cell of any one of claims 21 to 25, wherein the exogenous nucleic acid sequences comprise an extending packaging region (EPR) between the 5 ’ promoter and the selectable marker.

27. The host cell of claim 26, wherein the EPR comprises multiple potential Kozak sequences and / or ATG translation start sites.

28. The host cell of any one of claims 21 to 27, wherein the promoter sequence is selected from the group consisting of SIN-LTR, SV40, EFla, E. coli lac, E. coli trp, phage lambda PL, phage lambda PR, T3, T7, cytomegalovirus (CMV) immediate early, herpes simplex virus (HSV) thymidine kinase, alpha-lactalbumin, and mouse metallothionein-I promoter sequences.

29. The host cell of any one of claims 21 to 28, wherein the first promoter sequence is a weak promoter sequence.

30. The host cell of any one of claims 21 to 29, wherein the first promoter sequence is not a retroviral LTR promoter.

31. The host cell of any one of claims 21 to 30, wherein the integrated docking sites further comprise an exogenous promoter.

32. The host cell of claim 31, wherein the exogenous promoter is selected from the group consisting of SIN-LTR, SV40, EFla, E. coli lac, E. coli trp, phage lambda PL, phage lambda PR, T3, T7, cytomegalovirus (CMV) immediate early, herpes simplex virus (HSV) thymidine kinase, alpha-lactalbumin, and mouse metallothionein-I promoter sequences.

33. The host cell line of claim 32, wherein the promoter is a retroviral LTR.

34. The host cell line of claim 33, wherein the retroviral LTR is a SIN LTR.

35. The host cell of any one of claims 20 to 34, wherein the host cell line comprises a nucleic acid construct encoding an enzyme that facilitates insertion of the nucleic acid expression construct at the dock site.

36. The host cell of claim 35, wherein the nucleic acid construct encoding an enzyme that facilitates insertion of the exogenous nucleic acid sequences at the dock site is provided in a vector.

37. The host cell of claim 36, wherein the vector is a plasmid vector.

38. The host cell of any one of claims 35 to 37, wherein the enzyme is selected from the group consisting of an integrase, a recombinase, a nuclease and a nickase.

39. The host cell of any one of claims 20 to 38, wherein the host cell genome comprises from 5 to 500 integrated docking sites, each docking site comprising at least one dock site insertion element.

40. The host cell of any one of claims 20 to 29, wherein the integrated docking sites are independently positioned throughout the host cell genome.

41. The host cell of any one of claims 20 to 40, wherein the dock site insertion element is targeted by enzyme selected from the group consisting of an integrase, a recombinase, a nuclease and a nickase.

42. The host cell of any one of claims 20 to 41 , wherein the dock site insertion element is selected from the group consisting of a recombinase dock site insertion element and a HDR dock site insertion element.

43. The host cell of claim 42, wherein the dock site insertion element is a recombinase dock site insertion element.

44. The host cell of claim 43, wherein the recombinase dock site insertion element comprises an attachment site (att).

45. The host cell of claim 44, wherein the attachment site (att) is selected from the group consisting of attB and attP and attR and attL.

46. The host cell of claim 42, wherein the recombinase dock site insertion element comprises a LoxP sequence.

47. The host cell of claim 42, wherein the recombinase dock site insertion element is a Flp Recombination Target (FRT) site.

48. The host cell of claim 42, wherein the dock site insertion element is a HDR dock site insertion element.

49. The host cell of claim 48, wherein the HDR dock site insertion element comprises one or two dock site homology arms.

50. The host cell of claim 49, wherein the HDR dock site insertion element further comprises one or more sequences homologous to a guide RNA sequence.

51. The host cell of any of claims 48 to 50, wherein the dock site homology arms are from about 30 to 1000 bases in length.

52. The host cell of any one of claims 19 to 50, wherein each docking site is flanked by exogenous integrating vector sequences.

53. The host cell of claim 52, wherein the exogenous integrating vector sequences are selected from the group consisting of viral vector sequences and transposon vector sequences.

54. The host cell of any one of claims 20 to 53, wherein the docking sites each further comprise a sequence encoding a selectable maker operably linked to a promoter.

55. The host cell of any one of claims 20 to 54, wherein the dock site insertion element is positioned to facilitate cassette exchange.

56. The host cell of any one of claims 20 to 55, wherein each docking site comprises two dock site insertion elements.

57. The host cell of claim 56, wherein the two dock site insertion elements are positioned to facilitate cassette exchange.

58. The host cell of any one of claims 56 to 57, wherein the two dock site insertion elements flank sequences encoding a selectable marker, an enzyme, or a combination thereof.

59. The host cell of any one of claims 20 to 58, wherein the exogenous nucleic acid sequences further comprise a signal peptide sequence operably linked to the first product of interest.

60. The host cell of any one of claims 1 to 59, wherein the host cell is selected from the group consisting of Chinese Hamster Ovary (CHO) cells, HEK 293 cells, CAP cells, bovine mammary epithelial cells, monkey kidney CV1 line transformed by SV40, baby hamster kidney cells, mouse sertoli cells, monkey kidney cells, African green monkey kidney cells, human cervical carcinoma cells, canine kidney cells, buffalo rat liver cells, human lung cells, human liver cells, mouse mammary tumor, TRI cells, MRC 5 cells, FS4 cells, rat fibroblasts, MDBK cells and human hepatoma line cells.

61. The host cell line of claim 60, wherein the host cell is selected from the group consisting of a Chinese Hamster Ovary (CHO) cell, a HEK 293 cell and a CAP cell.

62. The host cell line of any one of claims 60 to 61 , wherein the host cell line is a GS knockout cell line.

63. The host cell line of any one of claims 60 to 61 , wherein the host cell line is a DHFR knockout cell line.

64. A cell culture comprising host cells of any of claims 1 to 63.

65. A method comprising: culturing a plurality of host cells according to any one of claims 1 to 62 in a culture medium under conditions such that the product of interest or second products of interests and / or an assembled complex thereof is produced; and purifying the product or products of interest or assembled complex thereof.

66. The method of claim 65, wherein the product or products of interest are secreted into the culture medium and purified therefrom.

67. A method comprising: introducing at least first nucleic acid constructs encoding a first protein or nucleic acid of interest and second nucleic acid constructs encoding a second protein or nucleic acid of interest at a ratio of first nucleic acid constructs to second nucleic acid constructs of from 1 : 1 to 5000:1 into a host cell having genome comprising from 1 to 500 integrated docking sites, each docking site comprising at least one dock site insertion element and the nucleic acid constructs each comprising at least one insertion element compatible with the at least one dock site insertion element in the integrated docking sites, under conditions such that the nucleic acid expression constructs are inserted at the dock sites at a ratio of first nucleic acid constructs to second nucleic acid constructs of at least 1:1.

68. The method of claim 67, wherein the ratio of inserted first nucleic acid constructs to second nucleic acid constructs is from 2:1 to 1000: 1.

69. The method of claim 67, wherein the ratio of inserted first nucleic acid constructs to second nucleic acid constructs is from 5: 1 to 500:1.

70. The method of claim 67, wherein the ratio of inserted first nucleic acid constructs to second nucleic acid constructs is from 10:1 to 200: 1.

71. The method of claim 67, wherein the ratio of inserted first nucleic acid constructs to second nucleic acid constructs is from 10: 1 to 100: 1.

72. The method of any one of claims 67 to 71, wherein the first and second proteins of interest are subunits of a multi-subunit protein.

73. The method of any one of claims 67 to 71, wherein the first and second proteins are subunits of a viral particle or the first and second nucleic acids are part of a viral genome.

74. The method of any one of claims 67 to 71, wherein one of the first and second proteins is an enzyme.

75. The method of any one of claims 67 to 74, wherein only first nucleic acid constructs and second nucleic acid constructs are introduced into the host cell.

76. The method of any one of claims 67 to 75, further comprising introducing a third nucleic acid construct encoding a third protein of interest under conditions such that the nucleic acid expression constructs are inserted at the dock sites at a ratio of first nucleic acid construct or second nucleic acid construct to third nucleic acid construct selected from the group consisting of at least 1:1, at least 2: 1, from 1: 1 to 5000:1, from 2:1 to 1000:1, from 5:1 to 500: 1, from 10: 1 to 200: 1, and from 10: 1 to 100:1.

77. The method of claim 76, further comprising introducing a fourth nucleic acid construct encoding a fourth protein of interest under conditions such that the nucleic acid expression constructs are inserted at the dock sites at a ratio of first nucleic acid construct, second nucleic acid construct, or third nucleic acid construct to the fourth nucleic construct selected from the group consisting of at least 1 :1, at least 2: 1, from 1:1 to 5000:1, from 2:1 to 1000:1, from 5: 1 to 500:1, from 10:1 to 200:1, and from 10:1 to 100:1.

78. The method of claim 77, further comprising introducing a fifth nucleic acid construct encoding a fifth protein of interest under conditions such that the nucleic acid expression constructs are inserted at the dock sites at a ratio of first nucleic acid construct, second nucleic acid construct, third nucleic acid construct or fourth nucleic construct to the fifth nucleic acid construct selected from the group consisting of at least 1 : 1 , at least 2: 1, from 1 : 1 to 5000:1, from 2:1 to 1000: 1, from 5: 1 to 500: 1, from 10:1 to 200: 1, and from 10: 1 to 100:1.

79. The method of any one of claims 67 to 78, wherein the at least first and second nucleic acid constructs further comprise at least the following elements in operable association in 5’ to 3’ order: an internal promoter sequence; a nucleic acid sequence encoding the first protein of interest or second protein that is operably linked to the internal promoter; and a poly A signal sequence.

80. The method of claim 79, wherein the at least first and second nucleic acid constructs comprise a selectable marker sequence.

81. The method of claim 80, wherein the at least first and second nucleic acid constructs comprise different selectable marker sequences.

82. The method of claim 79, wherein one of the first and second nucleic acid constructs comprises a selectable marker sequence and the other of the first and second nucleic acid constructs does not comprise a selectable marker sequence.

83. The method of any one of claims 80 to 81 , wherein the selectable marker sequences are 5 ’ to the internal promoter sequence and are operably linked to a 5 ’ promoter sequence.

84. The method of any one of claims 80 to 83, wherein the nucleic acid construct comprises an extending packaging region (EPR) between the 5 ’ promoter and the selectable marker.

85. The method of claim 84, wherein the EPR comprises multiple potential Kozak sequences and / or ATG translation start sites.

86. The method of any one of claims 79 to 85, wherein the promoter sequence is selected from the group consisting of SIN-LTR, SV40, EFla, E. coli lac, E. coli trp, phage lambda PL, phage lambda PR, T3, T7, cytomegalovirus (CMV) immediate early, herpes simplex virus (HSV) thymidine kinase, alpha-lactalbumin, and mouse metallothionein-I promoter sequences.

87. The method of any one of claims 79 to 86, wherein the first promoter sequence is a weak promoter sequence.

88. The method of any one of claims 79 to 87, wherein the first promoter sequence is not a retroviral LTR promoter.

89. The method of any one of claims 79 to 88, wherein the integrated docking sites further comprise an exogenous promoter.

90. The method of claim 89, wherein the exogenous promoter is selected from the group consisting of SIN-LTR, SV40, EFla, E. coli lac, E. coli trp, phage lambda PL, phage lambdaPR, T3, T7, cytomegalovirus (CMV) immediate early, herpes simplex virus (HSV) thymidine kinase, alpha-lactalbumin, and mouse metallothionein-I promoter sequences.

91. The method of claim 90, wherein the promoter is a retroviral LTR.

92. The method of claim 91 , wherein the retroviral LTR is a SIN LTR.

93. The method of any one of claims 67 to 92, wherein the nucleic acid expression constructs are provided in a vector.

94. The method of claim 93, wherein the vector is a plasmid vector.

95. The method of any one of claims 93 to 94, wherein the vector is transiently introduced into the host cell.

96. The method of any one of claims 67 to 95, wherein the host cell line comprises a nucleic acid construct encoding an enzyme that facilitates insertion of the nucleic acid expression construct at the dock site.

97. The method of claim 96, wherein the nucleic acid construct encoding an enzyme that facilitates insertion of the nucleic acid expression construct at the dock site is transiently introduced into the host cell.

98. The method of claims 96 or 97, wherein the nucleic acid construct encoding an enzyme that facilitates insertion of the nucleic acid expression construct at the dock site is provided in a vector.

99. The method of claim 98, wherein the vector is a plasmid vector.

100. The method of any one of claims 96 to 99, wherein the ratio of the ratio of the nucleic acid constructs encoding an enzyme that facilitates insertion of the nucleic acid expression construct at the dock site to the nucleic acid expression constructs encoding a first protein of interest that are transiently introduced into the host cell is from 1 : 1000 to 1 : 10.

101. The method of any one of claims 96 to 100, wherein the enzyme is selected from the group consisting of an integrase, a recombinase, a nuclease and a nickase.

102. The method of any one of claims 96 to 101 , wherein the nucleic acid construct encoding an enzyme that facilitates insertion of the nucleic acid expression construct at the dock site is provided in a vector.

103. The method of any one of claims 67 to 102, wherein the host cell genome comprises from 5 to 500 integrated docking sites, each docking site comprising at least one dock site insertion element.

104. The method of any one of claims 67 to 103, wherein the host cell genome comprises from 5 to 250 integrated docking sites, each docking site comprising at least one dock site insertion element.

105. The method of any one of claims 67 to 104, wherein the host cell genome comprises from 5 to 100 integrated docking sites, each docking site comprising at least one dock site insertion element.

106. The method of any one of claims 67 to 105, wherein the integrated docking sites are independently positioned throughout the host cell genome.

107. The method of any one of claims 67 to 106, wherein the dock site insertion element is targeted by enzyme selected from the group consisting of an integrase, a recombinase, a nuclease and a nickase.

108. The method of any one of claims 67 to 107, wherein the dock site insertion element is selected from the group consisting of a recombinase dock site insertion element and a HDR dock site insertion element.

109. The method of claim 108, wherein the dock site insertion element is a recombinase dock site insertion element.

110. The method of claim 109, wherein the recombinase dock site insertion element comprises an attachment site (att).

111. The method of claim 110, wherein the attachment site (att) is selected from the group consisting of attB and attP and attR and attL.

112. The method of claim 109, wherein the recombinase dock site insertion element comprises a LoxP sequence.

113. The method of claim 109, wherein the recombinase dock site insertion element is a Flp Recombination Target (FRT) site.

114. The method of claim 108, wherein the dock site insertion element is a HDR dock site insertion element.

115. The method of claim 114, wherein the HDR dock site insertion element comprises one or two dock site homology arms.

116. The method of claim 115, wherein the HDR dock site insertion element further comprises one or more sequences homologous to a guide RNA sequence.

117. The method of any of claims 115 to 116, wherein the dock site homology arms are from about 30 to 1000 bases in length.

118. The method of claim 117, wherein the integrase dock site insertion element comprises an AAVS1 safe harbor locus sequence.

119. The method of any one of claims 67 to 118, wherein each docking site is flanked by exogenous integrating vector sequences.

120. The method of claim 119, wherein the exogenous integrating vector sequences are selected from the group consisting of viral vector sequences and transposon vector sequences.

121. The method of any one of claims 67 to 120, wherein the docking sites each further comprise a sequence encoding a selectable maker operably linked to a promoter.

122. The method of any one of claims 67 to 121 , wherein the host cell further comprises an expression construct encoding an exogenous enzyme selected from the group consisting of an integrase, a recombinase, a nuclease and a nickase.

123. The method of claim 122, wherein the expression construct encoding an exogenous enzyme selected from the group consisting of an integrase, a recombinase, a nuclease and a nickase is provided in an episomal expression vector.

124. The method of claim 123, wherein the expression construct encoding an exogenous enzyme selected from the group consisting of an integrase, a recombinase, a nuclease and a nickase is integrated into the host cell genome.

125. The method of any one of claims 67 to 124, wherein the dock site insertion element is positioned to facilitate cassette exchange.

126. The method of any one of claims 67 to 125, wherein each docking site comprises two dock site insertion elements.

127. The method of claim 126, wherein the two dock site insertion elements are positioned to facilitate cassette exchange.

128. The method of any one of claims 126 to 127, wherein the two dock site insertion elements flank sequences encoding a selectable marker, an enzyme, or a combination thereof.

129. The method of any one of claims 126 to 127, wherein the nucleic acid expression constructs further comprise a signal peptide sequence operably linked to the first protein of interest.

130. The method of claim 129, wherein the signal peptide sequence is selected from the group consisting of tissue plasminogen activator, human growth hormone, lactoferrin, alphacasein and alpha-lactalbumin signal peptide sequences.

131. The method of any one of claims 67 to 130, wherein the nucleic acid expression constructs further comprise a protein purification marker sequence.

132. The method of claim 131, wherein the protein purification marker sequence is a hexahistidine tag or a hemagglutinin (HA) tag.

133. The method of any one of claims 67 to 132, wherein the host cell is selected from the group consisting of Chinese Hamster Ovary (CHO) cells, HEK 293 cells, CAP cells, bovine mammary epithelial cells, monkey kidney CV1 line transformed by SV40, baby hamster kidney cells, mouse sertoli cells, monkey kidney cells, African green monkey kidney cells, human cervical carcinoma cells, canine kidney cells, buffalo rat liver cells, human lung cells, human liver cells, mouse mammary tumor, TRI cells, MRC 5 cells, FS4 cells, rat fibroblasts, MDBK cells, VERO cells, NSO cells, and human hepatoma line cells.

134. The method of claim 133, wherein the host cell is selected from the group consisting of a Chinese Hamster Ovary (CHO) cell, a HEK 293 cell and a CAP cell.

135. The method of any one of claims 133 to 134, wherein the host cell is a GS knockout cell line.

136. The method of any one of claims 133 to 134, wherein the host cell is a DHFR knockout cell line.

137. A cell culture comprising host cells made the method on any of claims 67 to 136.

138. A process for producing a protein of interest comprising culturing host cells according to claim 137 under conditions that the protein of interest is expressed and purifying the protein of interest from the host cell culture.

139. The process of claim 138, wherein the host cells are grown in a medium comprising an inhibitor of the selectable marker.

140. The process of claim 139, wherein the selectable marker is GS and the inhibitor is phosphinothricin or methionine sulphoximine (Msx).

141. The process of claim 139, wherein the selectable marker is DHFR and the inhibitor is methotrexate.

142. A host cell comprising: a plurality of docking sites integrated into the genome of the host cell, each docking site comprising at least one dock site insertion element; at least integrated first nucleic acid constructs comprising at least one insertion element compatible with the dock site insertion element and encoding a first protein or nucleic acid of interest, and at least integrated second nucleic acid constructs comprising at least one insertion element compatible with the dock site insertion and encoding a second protein or nucleic acid of interest wherein the at least integrated first nucleic acid constructs and the at least second integrated nucleic acid constructs are integrated at the plurality of docking sites at a ratio of first nucleic acid constructs to second nucleic acid constructs of at least 1: 1.

143. The host cell of claim 142, wherein the ratio of integrated first nucleic acid constructs to second nucleic acid constructs is from 2:1 to 1000:1.

144. The host cell of claim 142, wherein the ratio of integrated first nucleic acid constructs to second nucleic acid constructs is from 5:1 to 500:1.

145. The host cell of claim 142, wherein the ratio of integrated first nucleic acid constructs to second nucleic acid constructs is from 10: 1 to 200: 1.

146. The host cell of claim 142, wherein the ratio of integrated first nucleic acid constructs to second nucleic acid constructs is from 10:1 to 100:1.

147. The host cell of any one of claims 142 to 146, wherein the first and second proteins of interest are subunits of a multi-subunit protein.

148. The host cell of any one of claims 142 to 146, wherein the first and second proteins are subunits of a viral particle or the first and second nucleic acids are part of a viral genome.

149. The host cell of any one of claims 142 to 146, wherein one of the first and second proteins is an enzyme.

150. The host cell of any one of claims 142 to 149, wherein only first nucleic acid constructs and second nucleic acid constructs are introduced into the host cell.

151. The host cell of any one of claims 142 to 150, further comprising a third nucleic acid construct encoding a third protein of interest at a ratio of first nucleic acid construct or second nucleic acid construct to third nucleic acid construct selected from the group consisting of at least 1:1, at least 2:1, from 2:1 to 1000:1, from 5:1 to 500: 1, from 10: 1 to 200:1, and from 10:1 to 100: 1.

152. The host cell of claim 151, further comprising a fourth nucleic acid construct encoding a fourth protein of interest at a ratio of first nucleic acid construct, second nucleic acid construct, or third nucleic acid construct to the fourth nucleic construct selected from the group consisting of at least 1 :1, at least 2:1, from 2:1 to 1000: 1, from 5: 1 to 500:1, from 10:1 to 200: 1, and from 10:1 to 100:1.

153. The host cell of claim 152, further comprising a fifth nucleic acid construct encoding a fifth protein of interest at a ratio of first nucleic acid construct, second nucleic acid construct, third nucleic acid construct or fourth nucleic construct to the fifth nucleic acid construct selected from the group consisting of at least 1:1, at least 2:1, from 2:1 to 1000:1, from 5:1 to 500:1, from 10:1 to 200:1, and from 10: 1 to 100:1.

154. The host cell of any one of claims 142 to 153, wherein the at least first and second nucleic acid constructs further comprise at least the following elements in operable association in 5’ to 3’ order: an internal promoter sequence; a nucleic acid sequence encoding the first protein of interest or second protein that is operably linked to the internal promoter; and a poly A signal sequence.

155. The host cell of claim 154, wherein the at least first and second nucleic acid constructs comprise a selectable marker sequence.

156. The host cell of claim 155, wherein the at least first and second nucleic acid constructs comprise different selectable marker sequences.

157. The host cell of claim 156, wherein one of the first and second nucleic acid constructs comprises a selectable marker sequence and the other of the first and second nucleic acid constructs does not comprise a selectable marker sequence.

158. The host cell of any one of claims 155 to 157, wherein the selectable marker sequences are 5’ to the internal promoter sequence and are operably linked to a 5’ promoter sequence.

159. The host cell of any one of claims 155 to 158, wherein the nucleic acid construct comprises an extending packaging region (EPR) between the 5 ’ promoter and the selectable marker.

160. The host cell of claim 159, wherein the EPR comprises multiple potential Kozak sequences and / or ATG translation start sites.

161. The host cell of any one of claims 154 to 160, wherein the promoter sequence is selected from the group consisting of SIN-LTR, SV40, EFla, E. coli lac, E. coli trp, phage lambda PL, phage lambda PR, T3, T7, cytomegalovirus (CMV) immediate early, herpes simplex virus (HSV) thymidine kinase, alpha-lactalbumin, and mouse metallothionein-I promoter sequences.

162. The host cell of any one of claims 154 to 161, wherein the first promoter sequence is a weak promoter sequence.

163. The host cell of any one of claims 154 to 162, wherein the first promoter sequence is not a retroviral LTR promoter.

164. The host cell of any one of claims 154 to 163, wherein the integrated docking sites further comprise an exogenous promoter.I l l165. The host cell of claim 164, wherein the exogenous promoter is selected from the group consisting of SIN-LTR, SV40, EFla, E. coll lac, E. coli trp, phage lambda PL, phage lambda PR, T3, T7, cytomegalovirus (CMV) immediate early, herpes simplex virus (HSV) thymidine kinase, alpha-lactalbumin, and mouse metallothionein-I promoter sequences.

166. The host cell of claim 165, wherein the promoter is a retroviral LTR.

167. The host cell of claim 166, wherein the retroviral LTR is a SIN LTR.

168. The host cell of any one of claims 142 to 167, wherein the nucleic acid expression constructs are provided in a vector.

169. The host cell of claim 168, wherein the vector is a plasmid vector.

170. The host cell of any one of claims 142 to 169, wherein the host cell comprises a nucleic acid construct encoding an enzyme that facilitates insertion of the nucleic acid expression construct at the dock site.

171. The host cell of claim 170, wherein the nucleic acid construct encoding an enzyme that facilitates insertion of the nucleic acid expression construct at the dock site is provided in a vector.

172. The host cell of claim 171 , wherein the vector is a plasmid vector.

173. The host cell of any one of claims 169 to 173, wherein the ratio of the ratio of the nucleic acid constructs encoding an enzyme that facilitates insertion of the nucleic acid expression construct at the dock site to the nucleic acid expression constructs encoding a first protein of interest that are transiently introduced into the host cell is from 1:1000 to 1:10.

174. The host cell of any one of claims 169 to 173, wherein the enzyme is selected from the group consisting of an integrase, a recombinase, a nuclease and a nickase.

175. The host cell of any one of claims 169 to 174, wherein the nucleic acid construct encoding an enzyme that facilitates insertion of the nucleic acid expression construct at the dock site is provided in a vector.

176. The host cell of any one of claims 142 to 175, wherein the host cell genome comprises from 5 to 500 integrated docking sites, each docking site comprising at least one dock site insertion element.

177. The host cell of any one of claims 142 to 176, wherein the host cell genome comprises from 5 to 250 integrated docking sites, each docking site comprising at least one dock site insertion element.

178. The host cell of any one of claims 142 to 177, wherein the host cell genome comprises from 5 to 100 integrated docking sites, each docking site comprising at least one dock site insertion element.

179. The host cell of any one of claims 142 to 178, wherein the integrated docking sites are independently positioned throughout the host cell genome.

180. The host cell of any one of claims 142 to 179, wherein the dock site insertion element is targeted by enzyme selected from the group consisting of an integrase, a recombinase, a nuclease and a nickase.

181. The host cell of any one of claims 142 to 180, wherein the dock site insertion element is selected from the group consisting of a recombinase dock site insertion element and a HDR dock site insertion element.

182. The host cell of claim 181, wherein the dock site insertion element is a recombinase dock site insertion element.

183. The host cell of claim 182, wherein the recombinase dock site insertion element comprises an attachment site (att).

184. The host cell of claim 183, wherein the attachment site (att) is selected from the group consisting of attB and attP and attR and attL.

185. The host cell of claim 182, wherein the recombinase dock site insertion element comprises a LoxP sequence.

186. The host cell of claim 182, wherein the recombinase dock site insertion element is a Flp Recombination Target (FRT) site.

187. The host cell of claim 181, wherein the dock site insertion element is a HDR dock site insertion element.

188. The host cell of claim 187, wherein the HDR dock site insertion element comprises one or two dock site homology arms.

189. The host cell of claim 188, wherein the HDR dock site insertion element further comprises one or more sequences homologous to a guide RNA sequence.

190. The host cell of any of claims 188 to 189, wherein the dock site homology arms are from about 30 to 1000 bases in length.

191. The host cell of claim 180, wherein the integrase dock site insertion element comprises an AAVS1 safe harbor locus sequence.

192. The host cell of any one of claims 142 to 191, wherein each docking site is flanked by exogenous integrating vector sequences.

193. The host cell of claim 192, wherein the exogenous integrating vector sequences are selected from the group consisting of viral vector sequences and transposon vector sequences.

194. The host cell of any one of claims 142 to 193, wherein the docking sites each further comprise a sequence encoding a selectable maker operably linked to a promoter.

195. The host cell of any one of claims 142 to 194, wherein the host cell further comprises an expression construct encoding an exogenous enzyme selected from the group consisting of an integrase, a recombinase, a nuclease and a nickase.

196. The host cell of claim 195, wherein the expression construct encoding an exogenous enzyme selected from the group consisting of an integrase, a recombinase, a nuclease and a nickase is provided in an episomal expression vector.

197. The host cell of claim 195, wherein the expression construct encoding an exogenous enzyme selected from the group consisting of an integrase, a recombinase, a nuclease and a nickase is integrated into the host cell genome.

198. The host cell of any one of claims 142 to 197, wherein the dock site insertion element is positioned to facilitate cassette exchange.

199. The host cell of any one of claims 142 to 198, wherein each docking site comprises two dock site insertion elements.

200. The host cell of claim 199, wherein the two dock site insertion elements are positioned to facilitate cassette exchange.

201. The host cell of any one of claims 199 to 200, wherein the two dock site insertion elements flank sequences encoding a selectable marker, an enzyme, or a combination thereof.

202. The host cell of any one of claims 142 to 201 , wherein the nucleic acid expression constructs further comprise a signal peptide sequence operably linked to the first protein of interest.

203. The host cell of claim 202, wherein the signal peptide sequence is selected from the group consisting of tissue plasminogen activator, human growth hormone, lactoferrin, alphacasein and alpha-lactalbumin signal peptide sequences.

204. The host cell of any one of claims 142 to 203, wherein the nucleic acid expression constructs further comprise a protein purification marker sequence.

205. The host cell of claim 204, wherein the protein purification marker sequence is a hexahistidine tag or a hemagglutinin (HA) tag.

206. The host cell of any one of claims 142 to 205, wherein the host cell is selected from the group consisting of Chinese Hamster Ovary (CHO) cells, HEK 293 cells, CAP cells, bovine mammary epithelial cells, monkey kidney CV1 line transformed by SV40, baby hamster kidney cells, mouse sertoli cells, monkey kidney cells, African green monkey kidney cells, human cervical carcinoma cells, canine kidney cells, buffalo rat liver cells, human lung cells, human liver cells, mouse mammary tumor, TRI cells, MRC 5 cells, FS4 cells, rat fibroblasts, MDBK cells, VERO cells, NSO cells, and human hepatoma line cells.

207. The host cell of claim 206, wherein the host cell is selected from the group consisting of a Chinese Hamster Ovary (CHO) cell, a HEK 293 cell and a CAP cell.

208. The host cell of any one of claims 206 to 207, wherein the host cell is a GS knockout cell line.

209. The host cell of any one of claims 206 to 207, wherein the host cell is a DHFR knockout cell line.

210. A cell culture comprising host cells of any of claims 142 to 209.

211. A process for producing a protein of interest comprising culturing host cells according to any of claims 142 to 209 under conditions that the protein(s) of interest are expressed and purifying the protein(s) of interest from the host cell culture.

212. The process of claim 211, wherein the host cells are grown in a medium comprising an inhibitor of the selectable marker.

213. The process of claim 211, wherein the selectable marker is GS and the inhibitor is phosphinothricin or methionine sulphoximine (Msx).

214. The process of claim 211, wherein the selectable marker is DHFR and the inhibitor is methotrexate.

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