Isolation of polymorphic mRNAs (cDNAs) by in-gel competitive reassociation.
In-gel competitive reassociation (IGCR) is a method for differential subtraction of polymorphic (RFLP) DNA fragments between two DNA samples of interest without probes or specific sequence information. Previously IGCR was used to enrich and isolate polymorphic genomic DNA fragments from mouse and human genomic DNA. We have modified the original IGCR procedures specifically for the isolation of polymorphic mRNAs in the form of cDNAs. Here we demonstrate that polymorphic mRNAs (cDNAs) between BALB/c and C57BL/6J mice that result from alterations at restriction sites and insertions-deletions are isolated with high efficiency by the IGCR procedure. A high proportion of the cDNAs was enriched by IGCR and 80% of the enriched clones were found to be actual RFLP fragments, indicating that the procedure is also applicable to direct screening for polymorphic mRNAs.
Gotoh
Koshichi
K
Human Gene 2, Kazusa DNA Research Institute, 2-6-7, Kazusa-Kamatari, Kisarazu, Chiba 292-0818, Japan. koshichi@kazusa.or.jp
Oishi
Michio
M
D000818:Animals; D003001:Cloning, Molecular / Q000379:methods*; D018076:DNA, Complementary / Q000235:genetics* / Q000302:isolation & purification*; D051379:Mice; D008807:Mice, Inbred BALB C; D008810:Mice, Inbred C57BL; D008969:Molecular Sequence Data; D012150:Polymorphism, Restriction Fragment Length; D012333:RNA, Messenger / Q000235:genetics* / Q000302:isolation & purification*
D018076:DNA, Complementary; D012333:RNA, Messenger