5 results about "Bacterial artificial chromosome" patented technology

The invention belongs to the technical field of genetic engineering, and particularly relates to a universal bacterial artificial chromosome recombinant virus transfer vector and a construction method thereof, the transfer vector comprises a BAC amplification vector with pUC19-loxp-gpt-IRES-EGFP as a basic skeleton and pUC19-BAC as a functional sequence; according to the universal bacterial artificial chromosome recombinant virus transfer vector and the construction method thereof, a drug screening target gene reading frame and an EGFP fluorescent protein gene reading frame are connected through an IRES double-expression connection sequence, and double screening can be carried out; in the BAC construction process, screening genes and functional sequences are respectively constructed to a high-copy T vector, so that sufficient plasmid DNA can be conveniently copied and prepared. And through a proper restriction enzyme cutting site, introduction of various homologous arms is facilitated, the alignment is wide, simultaneous operation of various genomic sequences is facilitated, and the research efficiency is improved.

The disclosure provides for methods of constructing synthetic chromosomes including the steps of providing host cells with an endogenous chromosome, transforming a cloning vector and a cloning cassette into the host cells, excising target genomic nucleic acids from the endogenous chromosome, recombining the excised target genomic nucleic acids with the cloning cassette via homologous recombination to form heterologous vectors comprising cloned sequences, extracting the heterologous vectors containing the cloned sequences from the host cells, digesting the heterologous vectors with a restriction endonuclease to release the cloned sequences from the heterologous vectors to provide released cloned sequences, and introducing the released cloned sequences, a centromere cassette, and a yeast artificial chromosomes or bacterial artificial chromosomes into a second host cell, such that the released cloned sequences, the centromere cassette, and the YAC or BAC recombine with one another via homologous recombination to produce the synthetic chromosome.

This invention provides a method for seamless editing of a double-copy target gene in a bacterial artificial chromosome. The method includes: first, constructing a bacterial artificial chromosome containing a double-copy target gene and introducing it into *E. coli* SW102 to obtain a first recombinant bacterium; second, constructing a target fragment including a chloramphenicol resistance gene, a gentamicin resistance gene, and a galactokinase encoding gene, contacting this target fragment with the first recombinant bacterium, and obtaining a second recombinant bacterium through a first homologous recombination; finally, constructing a target fragment for editing the target gene and obtaining a third recombinant bacterium through a second homologous recombination, thus completing the seamless editing of the double-copy gene. The method provided by this invention can simultaneously recombine and modify a double-copy target gene in a bacterial artificial chromosome, avoiding the problems of low double-site knock-in rate and high screening difficulty associated with single galactokinase encoding gene marker screening, and achieving seamless editing of the double-copy target gene.

The present invention relates to a process for preparing recombinant Modified Vaccina Virus Ankara (MVA) using bacterial artificial chromosome (BAC) based permanent dominant selection.

This invention discloses a novel antitumor macrolide compound, UCN370, its preparation method, and its applications. The structural formula of compound UCN370 is shown below. This invention successfully expressed the bacterial artificial chromosome (BAC) plasmid pUCN370, containing the UCN370 biosynthetic gene cluster, using the chassis strain *Streptomyces albus* J1074, resulting in the novel scaffold compound UCN370. The novel antitumor macrolide compound UCN370 described in this invention exhibits potent inhibitory activity against tumor cells such as HK-2 and can be used to prepare antitumor drugs.