Sequence-independent detection of gene family homologs: identification of a transcript encoding a molluscan serine protease homologous to the pancreatic enzymes of vertebrates.
Autoradiography of 32P-labeled cDNA, fractionated at high resolution by electrophoresis through thin (0.8-1.5 mm) vertical alkaline agarose gels, provides a sequence-independent screening procedure for gene family homologs. A screen of tissues of a marine mollusc revealed a prominent intestine-specific cDNA encoding a pancreatic serine protease homolog, which was not detectable as a discrete poly(A)+ RNA species on formaldehyde agarose gels. Discrete cDNA products are authentic, non-truncated transcripts of tissue-specific mRNA. A band-sharpening effect is imparted to cDNA products due to (a) substitution of a uniform length 5'-oligo(dT) terminus for heterogeneous 3'-poly(A) termini and (b) the inherent superior resolution of alkaline-denatured DNA.
Groppe
J C
JC
Marine Biotechnology Center, University of California, Santa Barbara 93106.
Morse
D E
DE
D000818:Animals; D015152:Blotting, Northern; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000737:chemistry / Q000302:isolation & purification; D015723:Gene Library; D005880:Gills / Q000737:chemistry; D006863:Hydrogen-Ion Concentration; D007422:Intestines / Q000737:chemistry; D008974:Mollusca / Q000201:enzymology / Q000235:genetics*; D009691:Nucleic Acid Denaturation; D010179:Pancreas / Q000201:enzymology*; D012333:RNA, Messenger / Q000032:analysis*; D017422:Sequence Analysis, DNA; D017385:Sequence Homology; D012697:Serine Endopeptidases / Q000235:genetics*
D018076:DNA, Complementary; D012333:RNA, Messenger; D012697:Serine Endopeptidases