[Reconstruction of large genomic segment coding for human immunoglobulin kappa chain by meiotic homologous recombination of yeast artificial chromosome].
Meiotic homologous recombination between overlapping yeast artificial chromosome (YAC) can be used to reconstruct larger genomic segment in vivo. We have screened CEPH and Olson human genomic YAC libraries and obtained two YAC clones containing the large genomic segments coding for partial human Ig kappa. In order to reconstruct whole gene cluster of human Ig kappa, two yeast cells containing the different YAC described over were mated and induced to undergo meiotic division and sporulation. Analyzing the results of PCR and Southern blotting, we obtained a clone containing a single recombinant YAC with 400 kb in length, which spans almost the entire gene cluster of human Ig kappa, including 32 V kappa, 5 J kappa, constant domain genes. Besides, we have improved and simplified the traditional method of meiotic recombination between overlapping YACs, and the results indicate the feasibility of this method to reconstruct any larger intact genomic segments.
Nie
Zhi Yan
ZY
Institute of Biochemistry and Cell Biology, Shanghai Institute for Biological Science, Chinese Academy of Sciences, Shanghai Cel-star Institude of Biotechnology, Shanghai.
Lu
Bu Feng
BF
Xu
Yan Ping
YP
Lu
Bo
B
Guo
Li He
LH
D018244:Chromosomes, Artificial, Yeast / Q000235:genetics*; D003001:Cloning, Molecular; D015698:Genomic Library; D006801:Humans; D007145:Immunoglobulin kappa-Chains / Q000235:genetics*; D008540:Meiosis; D011995:Recombination, Genetic
D007145:Immunoglobulin kappa-Chains