Single Nucleotide Polymorphism-based Identification of Bacterial Artificial Chromosome-mediated Homologous Recombination

被引:0
|
作者
Chung, Sun-Ku [1 ,2 ]
机构
[1] Korea Inst Oriental Med, Div Korean Med KM Sci Res, Daejeon 34054, South Korea
[2] Univ Sci & Technol, Korea Inst Oriental Med KIOM Campus, Korean Convergence Med Sci, Daejeon 34054, South Korea
来源
FRONTIERS IN BIOSCIENCE-LANDMARK | 2024年 / 29卷 / 08期
关键词
single nucleotide polymorphism; bacterial artificial chromosome; homologous recombination; single-stranded conformational polymorphism; restriction fragment length polymorphism; human pluripotent stem cell; PLURIPOTENT STEM-CELLS; TRANSGENIC MICE; ESCHERICHIA-COLI; SNP PANEL; DISEASE; EXPRESSION; INTEGRATION; TALEN; RISK; ZFN;
D O I
10.31083/j.fbl2908280
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Bacterial Artificial chromosome (BAC) recombineering is a powerful genetic manipulation tool for the efficient development of recombinant genetic resources. Long homology arms of more than 150 kb composed of BAC constructs not only substantially enhance genetic recombination events, but also provide a variety of single nucleotide polymorphisms (SNPs) that are useful markers for accurately docking BAC constructs at target sites. Even if the BAC construct is homologous to the sequences of the target region, different variations may be distributed between various SNPs within the region and those within the BAC construct. Once the BAC construct carrying these variations was precisely replaced in the target region, the SNP profiles within the target genomic locus were directly replaced with those in the BAC. This alteration in SNP profiles ensured that the BAC construct accurately targeted the designated site. In this study, we introduced restriction fragment length polymorphism or single-strand conformation polymorphism analyses to validate and evaluate BAC recombination based on changes in SNP patterns. These methods provide a simple and economical solution to validation steps that can be cumbersome with large homologous sequences, facilitating access to the production of therapeutic resources or disease models based on BAC-mediated homologous recombination.
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页数:11
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