[Enzymatic synthesis and molecular cloning of the pigeon alpha-globin structural gene].

Double-stranded DNA synthesized from the pigeon globin mRNA by the subsequent actions of avian myeloblastosis virus reverse transcriptase and E. coli DNA polymerase I was split with nuclease S1 and inserted into PstI site in the plasmid pBR322 by poly(dG) times poly(dC) homopolymer extension technique using terminal deoxynucleotidyl transferase. E. coli transformants have been shown to contain pigeon globin sequences by colony hybridization with pigeon globin [32P]cDNA. The inserted DNA fragment deleted from recombinant DNA by PstI treatment hybridizes with globin cDNA. The maximal length of the inserted fragment measured in agarose gel was found to be 550--560 base pairs. Inserted sequences subjected to analysis by hybridization with alpha- and beta-[32P]cDNA have been ascribed to the pigeon alpha globin chain. EcoRI, HindIII, BglII, SalI, BamHI, PstI restriction enzymes did not cleave the inserted DNA fragment. Pigeon DNA coding alpha-globin chain contains recognition sites for AluI, HindII and HaeIII restriction enzymes. Part of the recombinant clones remains resistant to ampicillin and therefore in some of these clones the globin gene could be expressed.