HBD3 mammary gland specific expression vector and constructed recombinant cell

Abstract

The invention discloses a HBD3 mammary gland specific expression vector and a constructed recombinant cell. The vector comprises a HBD3 gene with a signal transduction peptide, and CSN5 and CSN3 are set upstream and downstream of the HBD3 gene respectively for serving as regulating and controlling elements. The host cell of the recombinant cell is a bovine fetal fibroblast cell, an exogenous expression vector pARNG-HBD3 is integrated to a pseudo attP locus under the action of phiC31 integrase, positive cloning is realized through medicament screening, and an antibiotic screening marker between two homodromous LoxP sequences on the vector is removed through Cre recombinase treatment. A HBD3-containing bovine fetal fibroblast cell from which an antibiotic screening marker is removed is taken as a nuclear donor cell of a transgenic cloning embryo through nucleus transplantation, gene cloning embryos are obtained through SCNT (Somatic Cell Nuclear Transfer), and a milk cow which is free from a HBD3 transgenic gene of the antibiotic screening marker can be produced by transferring these embryos into a receptor cow.

Description

technical field [0001] The invention belongs to the field of genetic engineering and relates to a site-specific integrated expression vector, in particular to a HBD3 mammary gland-specific expression vector and constructed recombinant cells. Background technique [0002] Mastitis is a very common disease in dairy cows. This inflammation is caused by a variety of pathogenic microorganisms, mainly Escherichia coli (E.coli) and Staphylococcus (S.aureus) (Bannerman DD, Paape MJ, Lee JW, Zhao X, Hope JC, Rainard P. Escherichia coli and Staphylococcus aureus elicit differential innate immune responses following intramammary infection. Clin Diagn Lab Immunol. 2004 May; 11(3): 463-72.). Mastitis has caused huge economic losses to the dairy industry. At present, the treatment of mastitis is still dominated by the use of antibiotics, but antibiotics are expensive; and long-term use of antibiotics will cause bacteria to develop drug resistance, so a new treatment for mastitis is sought...

AI Technical Summary

Problems solved by technology

However, the selection of marker genes also brings some disadvantages: there are only a few effective marker genes widely used at present, and it is often necessary to perform multiple transformations on the same gene fragment during genetic engineering operations, and it is difficult to find suitable selection markers at this time. genes; in addition, the expression of selectable marker genes may have negative effects on the expression of other genes in transformed cells, leading to complicating the analysis of results; at the same time, the presence of selectable marker genes in transgenic crops increases the uncertainty of their distribution environment, This brings about environmental safety issues of transgenic animals and plants, which have a great restrictive effect on the environmental release of transgenic animals and plants

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