704 ISOLATION OF DNA SEGMENTS FROM THE HUMAN X CHROMOSOME

Abstract

We have utilized the methods of Gusella et al (PNAS 77, 2829, 1980) to isolate DNA segments from the human X chromosome. DNA was prepared from a human-Chinese hamster hybrid cell line containing the human X and essentially no other human chromosome. This DNA was ligated to phage DNA, the recombinant phage encapsulated and grown in E.coli. The plaques were exposed to labelled whole human and Chinese hamster DNA. Plaques which did not anneal with Chinese hamster DNA but did anneal with human DNA were isolated. The human DNA segments were recovered from these selected recombinant phage and tested for their ability to anneal with DNA from hybrid cells containing various human chromosomes. Six different recombinant phage have been studied sufficiently to demonstrate that they contain human DNA which is complementary to the human X and apparently to no other human chromosome. Further studies with hybrid cells containing known portions of the human X indicate that the recombinant DNA segments are derived from specific regions of the X. The methods utilized are efficient and demonstrate the feasibility of isolating a large number of DNA segments from the human X. These segments are suitable for detecting DNA polymorphisms and for study of X chromosome inactivation. (Presented in part to the Association for Research in Nervous and Mental Disease, December 5, 1980.)