Synthesis of DNA complementary to separated human alpha and beta globin messenger RNAs.
Human globulin messenger RNA, purified by oligo(dT)-cellulose column chromatography, is reproducibly separated into two bands by polyacrylamide gel electrophoresis in the presence of 99% formamide. The more rapidly migrating (fast) band is somewhat more abundant than the slow band in normal (nonthalassemic) total reticulocyte globin messenger RNA. In alpha-thalassemic (Hb H disease) messenger RNA, the slow band is 6.5 times more abundant than the fast band, whereas in beta-thalassemic messenger RNA the fast band is three times more abundant than a second band, which has a slightly greater mobility than the slow band of normal and alpha-thalassemic RNA. The RNA bands of nonthalassemic globin messenger RNA were eluted from the gel and efficiently transcribed into DNA copies by use of the RNA-dependent DAN polymerase of avian myeloblastosis virus. Hybridization of these copy DNAs to fast and slow band RANs and to nonfractionated normal, alpha-thalassemic, and geta-thalassemic messenger RNAs revealed that the eluted fast band RNA contains predominantly alpha-chain specific sequences, whereas the eluted slow band RNA contains predominantly beta-chain specific sequences. Nucleotide sequence analysis of 32-P-labeled RNA transcribed from the slow band copy DNA also indicated that the slow band RNA is beta messenger RNA.
D009189:Avian Myeloblastosis Virus / Q000201:enzymology; D001483:Base Sequence; D004247:DNA / Q000096:biosynthesis*; D004591:Electrophoresis, Polyacrylamide Gel; D005914:Globins; D006453:Hemoglobinopathies / Q000097:blood; D009693:Nucleic Acid Hybridization; D012333:RNA, Messenger / Q000032:analysis / Q000302:isolation & purification / Q000378:metabolism*; D012194:RNA-Directed DNA Polymerase / Q000378:metabolism; D012156:Reticulocytes / Q000032:analysis; D013789:Thalassemia / Q000097:blood; D014158:Transcription, Genetic
D012333:RNA, Messenger; D005914:Globins; D004247:DNA; D012194:RNA-Directed DNA Polymerase