ABSTRACT
We have examined 25 independent rat genomic clones each of which contains a U1 RNA gene or a pseudogene. We have found five clones whose restriction maps are identical with or overlapping one another. These clones contain sequences which are co-linear with that of U1 RNA except for one or two nucleotides (nt). They also contain 21-23-nt poly(A) stretches immediately after U1 RNA homology. In addition, the U1 RNA-poly(A) regions are surrounded by the same direct repeat sequences. Therefore, they seem to be pseudogenes which have been generated by the reverse transcription of poly(A)-tailed U1 RNA followed by the insertion of the transcript into the genome. Furthermore, conservation of the sequences extends over at least 18 kb of the flanking sequences. This suggests a family of conserved reverse-transcribed pseudogenes, which implies amplification of an original pseudogene. It is also suggested that the target sequence without insertion of a U1 RNA sequence has been amplified. The mechanisms for the amplification and sequence conservation are discussed.
Subject(s)
Gene Amplification , Genes , RNA, Small Nuclear/genetics , Transcription, Genetic , Animals , Base Sequence , Cloning, Molecular , DNA/isolation & purification , DNA Restriction Enzymes , Liver/metabolism , Rats , Rats, Inbred Strains , Sequence Homology, Nucleic AcidABSTRACT
Six phage clones that contain sequences hybridizable with the small nuclear RNA U2 were isolated from a rat gene library. Of these clones, one which includes a candidate for a functional U2 RNA gene was selected and characterized. The sequence within the clone which hybridizes with rat U2 RNA was completely co-linear with that of the RNA. A T-A-T-A box was not found in the region of more than 400 base-pairs which lies upstream of the gene. However, several block homologies were found with the upstream sequences of a rat U1 RNA gene candidate cloned in our laboratory. An "identifier sequence", which was reported to be an element of gene regulation related to differentiation, was found downstream of the coding region at the same distance and with the same orientation as the identifier sequence located downstream of the U1 RNA gene candidate. We detected a presumed U2 RNA precursor elongated by about 11 nucleotides at the 3' end by S1 nuclease mapping using a fragment from the clone. A potential termination signal for transcription was found within the elongated region of the presumed precursor. Southern blot analysis suggests that families of U2 RNA genes that have conserved flanking sequences are present in the genomes of rat, mouse, man, calf and chicken.
Subject(s)
Cloning, Molecular , Genes , RNA/genetics , Animals , Base Sequence , Cattle , Chickens , DNA/analysis , DNA Restriction Enzymes , Humans , Mice , Nucleic Acid Hybridization , RNA/analysis , RNA, Small Nuclear , Rats , Ribonuclease T1/metabolismABSTRACT
Four phage clones which hybridize with U1 small nuclear RNA were obtained from a rat gene library. Two clones contain a presumed pseudogene. A third clone includes two gene candidates that are co-linear with the rat U1-RNA, 3.6kb apart and in the opposite orientation. The two genes are surrounded by identical sequences of 491bp upstream and 178bp downstream. The upstream sequences do not contain a TATA box, but share many block homologies with those for the human U1-RNA gene(1-3). A 101bp "identifier (ID) sequence", which was reported to be specifically expressed in rat brain (4), is inserted immediately after the shared sequence downstream of one of the genes. In the fourth clone, there are two putative pseudogenes, which have one or three nucleotide changes, 3kb apart and in the same orientation. Southern blot analysis of total rat DNA reveals about 50 U1-RNA genes/pseudogenes in the genome.
