Purification of major abundance class of poly(A+)-RNA from rat ventral prostate.
Poly(A+)--RNA from rat ventral prostate was isolated using oligo(dT)-cellulose chromatography. 45% of the total poly(A+)--RNA was a single peak at 10S as demonstrated by centrifugation in a 5-20% sucrose gradient containing 1% SDS. By using complementary DNA probes, it was shown that the 10S RNA contained the major abundance class of poly(A+)--RNA. Denaturing agarose-gel analysis revealed 2 major bands in the 10S poly(A+)--RNA preparation approx. 600 NT and 500 NT (NT = nucleotides) long, resp. Double-stranded 32 P-DNAs complementary to light side and heavy side of the 10S poly(A+)--RNA peak were synthesized and isolated using reverse transcriptase and hydroxyapatite (HAP) chromatography. Approx. 40% of the first strand of the cDNAs were converted to double-stranded structures with a Tm of 88 degrees C. HAP purified double-stranded material was 92% resistant to S1 nuclease. the DNA--DNA reannealing profile of double stranded 32 P-cDNA enriched for the 500 NT band gave a Cot 1/2 of approximately 7 X 10(-4) moles X sec X 1(-1) indicating a complexity for this enriched synthetic gene of 500-600 nucleotide pairs (NTP).
D000818:Animals; D004247:DNA / Q000032:analysis; D004262:DNA Restriction Enzymes / Q000032:analysis; D008297:Male; D009691:Nucleic Acid Denaturation; D009693:Nucleic Acid Hybridization; D011061:Poly A / Q000302:isolation & purification*; D011467:Prostate / Q000032:analysis; D012313:RNA / Q000302:isolation & purification*; D012333:RNA, Messenger / Q000302:isolation & purification*; D051381:Rats
D012333:RNA, Messenger; D011061:Poly A; D012313:RNA; D004247:DNA; D004262:DNA Restriction Enzymes