Structure of the constant and 3' untranslated regions of the murine Balb/c gamma 2a heavy chain messenger RNA.

The complete sequence for the constant and 3' untranslated regions of a mouse gamma 2a immunoglobulin heavy chain mRNA is reported. The sequence is 1093 nucleotides long coding for the CH1 (amino-acids 118-214), the Hinge (215-230), the CH2 (231-340) and the CH3 (341-447). The 3' untranslated region is 103 nucleotides long preceding the poly(A). The nucleotide sequence predicts as in the case for gamma 1 and gamma 2b heavy chains an additional lysine residue before the termination codon. This sequence has been compared to the corresponding sequences of gamma 1 and gamma 2b heavy chain mRNAs. These sequences are respectively 75% and 84% homologous. The CH2 domains of gamma 2a and gamma 2b are 95% homologous at the nucleotide level. The cross-over point of a gamma 2a - gamma 2b heavy chain variant is located in a segment of 73 perfectly matching nucleotides. The 3' non coding regions of gamma 2a and gamma 2b are 89% homologous.     

The sequence of a cloned cDNA for the beta subunit of bovine thyrotropin predicts a protein containing both NH2-and COOH-terminal extensions

Cloned cDNAs containing sequences coding for the beta subunit of bovine thyrotropin have been identified. The complete nucleotide sequence of the largest of the beta subunit cDNA inserts has been determined. This cDNA contains 35 nucleotides from the 5' untranslated region of thyrotropin beta subunit mRNA and 60 nucleotides coding for an NH2-terminal precursor segment. This is followed by 339 nucleotides which code for the published amino acid sequence of the thyrotropin beta subunit. Following the 339 nucleotide beta subunit coding sequence, no termination codon is encountered for another 15 nucleotides. Thus, the cDNA codes for a thyrotropin beta subunit containing an additional 5 amino acids at the COOH terminus. The cDNA also contains 82 nucleotides of 3' untranslated sequence followed by a short poly(A) segment. Comparison of the bovine cDNA sequence to the recently described mouse thyrotropin beta subunit cDNA sequence reveals considerable homology throughout the coding sequence, including the COOH-terminal extension. These findings suggest the possibility that a thyrotropin beta subunit precursor is processed at both the NH2 and COOH termini.     

Sequence of the cDNA and gene for angiogenin, a human angiogenesis factor

Human cDNAs coding for angiogenin, a human tumor derived angiogenesis factor, were isolated from a cDNA library prepared from human liver poly(A) mRNA employing a synthetic oligonucleotide as a hybridization probe. The largest cDNA insert (697 base pairs) contained a short 5'-noncoding sequence followed by a sequence coding for a signal peptide of 24 (or 22) amino acids, 369 nucleotides coding for the mature protein of 123 amino acids, a stop codon, a 3'-noncoding sequence of 175 nucleotides, and a poly(A) tail. The gene coding for human angiogenin was then isolated from a genomic lambda Charon 4A bacteriophage library employing the cDNA as a probe. The nucleotide sequence of the gene and the adjacent 5'- and 3'-flanking regions (4688 base pairs) was then determined. The coding and 3'-noncoding regions of the gene for human angiogenin were found to be free of introns, and the DNA sequence for the gene agreed well with that of the cDNA. The gene contained a potential TATA box in the 5' end in addition to two Alu repetitive sequences immediately flanking the 5' and 3' ends of the gene. The third Alu sequence was also found about 500 nucleotides downstream from the Alu sequence at the 3' end of the gene. The amino acid sequence of human angiogenin as predicted from the gene sequence was in complete agreement with that determined by amino acid sequence analysis. It is about 35% homologous with human pancreatic ribonuclease, and the amino acid residues that are essential for the activity of ribonuclease are also conserved in angiogenin. This provocative finding is thought to have important physiological implications.     

Molecular cloning of bovine beta-lactoglobulin cDNA

A cDNA library from bovine mammary gland mRNA was constructed in pBR322 and screened by hybrid-selected translation and immunoscreening. Several beta-lactoglobulin clones were identified and sequenced. All clones contained cDNA fragments corresponding to the 3' region of beta-lactoglobulin mRNA. The 3' non-translated region of beta-lactoglobulin mRNA consists of 187 nucleotides; the polyadenylation signal AATAAA occurs 17 nucleotides before the poly(A) tail. The amino-acid sequence predicted from the 3' coding region corresponds completely to the previously determined amino-acid sequence of beta-lactoglobulin.     

Sequence of human adenosine deaminase cDNA including the coding region and a small intron

The nucleotide sequence for an unusual, cloned human adenosine deaminase cDNA has been determined. Contained within a sequence of 1535 nucleotides is a coding sequence of 1089 nucleotides that encodes a protein of 40,762 daltons. The coding sequence is interrupted by a non-coding region containing 76 nucleotides. Both the 3' and 5' ends of this region have consensus sequences generally associated with splice sites. The 3' untranslated sequence contained 308 nucleotides, including a polyadenylation signal sequence 20 nucleotides from the end. The cloned cDNA appears to correspond to a nuclear mRNA precursor which contains a small intron.     

Complete sequence of ovine alpha s2-casein messenger RNA

The primary structure of mRNA coding for ovine alpha s2 casein has been determined by chemical sequencing of three cDNA clones and the primer extension products of the longest one. The mRNA was 1,024 nucleotides long, excluding the poly(A) tail. The length of the 5' noncoding, coding and 3' noncoding regions was 53, 669 and 302 nucleotides, respectively. A comparison of the nucleotide sequence of ovine alpha s2-casein and guinea-pig casein A mRNAs revealed an extensive homology in the 5' and 3' noncoding regions. The deduced amino acid sequence of ovine alpha s2-casein was compared with its bovine and guinea-pig counterparts. Moreover, an heterogeneity was evidenced in the mRNA population of the alpha s2-casein.     

A mouse immunoglobulin heavy chain deletion mutant: isolation of a cDNA clone and sequence analysis of the mRNA

The mouse cell line IF2 secretes an immunoglobulin heavy chain lacking the CH1 domain. We have isolated and characterised a recombinant plasmid containing cDNA copies of the IF2 mutant mRNA. The cloned sequence extends from the nucleotides coding for amino acid 96 in the variable region through 100 nucleotides of untranslated region at the 3' end. The sequence of the cDNA insert reveals no discontinuity at the variable-hinge region junction, the site of the CH1 deletion. Experiments employing direct priming on the poly(A) tail of the IF2 heavy chain mRNA suggest that the 3' end of the cDNA clone (sequence C-C-C-T-G-C) is also the 3' end of the mRNA.     

Complete nucleotide sequence of ovine alpha-lactalbumin mRNA

The nucleotide sequence of ovine alpha-lactalbumin mRNA has been determined by chemical sequencing of two cDNA recombinant plasmids and a primer extension product. Ovine alpha-lactalbumin mRNA contains 723 nucleotides (excluding the poly(A) tail), with a 5' non-coding region of 26 nucleotides, followed by the 426 nucleotides of the coding region which determines a sequence signal of 19 amino acid residues and the 123 amino acid residues of mature alpha-lactalbumin. The coding region is followed by a 3' untranslated sequence of 271 nucleotides. The derived amino acid sequence of ovine pre-alpha-lactalbumin differs from that of its bovine counterpart by 8 amino acid substitutions, all but one originating from single mutations. Comparison of sequences of guinea pig, rat and human alpha-lactalbumin mRNAs with their ovine and bovine counterparts has revealed that these molecules have rapidly evolved. The highest degree of conservation was observed in the region coding for the mature protein and corresponds essentially to sequences which interact with UDP-galactosyltransferase and Ca2+ ions.     

Primary structure of Bacillus subtilis enolase gene deduced from c-DNA sequence

The nucleotide sequence for enolase gene of Bacillus subtilis was determined from recombinant clone pRE. The sequence was composed of 1570 bp which included the 1374 bp of the complete coding region, the 86 bp of the 5' noncoding region and the 110 bp of the 3' noncoding region containing a polyadenylation signal. In addition, the poly(A) tail was also found. A potential ribosome binding site was located 20 nucleotides upstream of the initiation codon in the 5' noncoding region. The aminoacid sequence deduced from the nucleotide sequence was 558 aminoacids in length. The size of the mRNA was 1.5 kb by the northern transfer technique.     

Isolation and nucleotide sequence of a full-length cDNA coding for aldolase B from human liver.

Two recombinant clones, pA2 and pA3, containing cDNA sequences for human aldolase B have been isolated from a full length human liver cDNA library. The larger one, pA3, has been subcloned in M13 phage and completely sequenced with the chain terminator method. The sequence covers 1,600 nucleotides including the whole coding region (1,050 nucleotides), 67 nucleotides from the 5' non-coding region and the whole 3' non-coding region, 440 nucleotides long, down to the poly-A tail. Comparison with rabbit aldolase A and with a partial sequence of rat aldolase B, shows a homology of about 76% for aldolase A and of about 94% for aldolase B, which indicates that the sequenced cDNA codes for the liver isoenzyme. This is the first complete sequence reported for human aldolase B. The pA3 clone strongly hybridizes to 18S mRNA from human adult liver as expected from the size of the isolated cDNA.     

Sequence of the yeast iso-1-cytochrome c mRNA

The nucleotide sequence of the yeast iso-1-cytochrome c (CYC1) mRNA is presented. The mRNA was enriched by hybridization to cloned CYC1 DNA attached to a solid matrix: either nitrocellulose filters or diazobenzyloxymethyl cellulose powder. The sequence of the 5'-end of the mRNA was determined by the extension of a CYC1-specific dodecanucleotide primer; the sequence of the 3'-end was determined using a decanucleotide d(pT8-G-A) primer. The CYC1 mRNA begins 61 nucleotides 5' to the AUG initiation codon, extends through the coding sequence to 172 to 175 nucleotides 3' to the UAA termination codon, followed by the poly(A) tail. There are no intervening sequences. Some of the sequences that the CYC1 mRNA shares in common with other eukaryotic mRNAs are discussed.     

The nucleotide sequence of cowpea mosaic virus B RNA

The complete sequence of the bottom component RNA (B RNA) of cowpea mosaic virus (CPMV) has been determined. Restriction enzyme fragments of double-stranded cDNA were cloned in M13 and the sequence of the inserts was determined by a combination of enzymatic and chemical sequencing techniques. Additional sequence information was obtained by primed synthesis on first strand cDNA. The complete sequence deduced is 5889 nucleotides long excluding the 3' poly(A), and contains an open reading frame sufficient to code for a polypeptide of mol. wt. 207 760. The coding region is flanked by a 5' leader sequence of 206 nucleotides and a 3' non-coding region of 82 residues which does not contain a polyadenylation signal.     

Molecular structure and flanking nucleotide sequences of the natural chicken ovomucoid gene

Five independent clones containing the natural chicken ovomucoid gene have been isolated from a chicken gene library. One of these clones, CL21, contains the complete ovomucoid gene and includes more than 3 kb of DNA sequences flanking both termini of the gene. Restriction endonuclease mapping, electron microscopy and direct DNA sequencing analyses of this clone have revealed that the ovomucoid gene is 5.6 kb long and codes for a messenger RNA of 821 nucleotides. The structural gene sequence coding Ifor the mature messenger RNA is split into at least eight segments by a minimum of seven intervening sequences of various sizes. The shortest structural gene segment is only 20 nucleotides long. All seven intervening sequences are located within the peptide coding region of the gene, and the sequences at the 5' and 3' untranslated regions of the mRNA are not interrupted by intervening sequences. The DNA sequences of the regions flanking the 5' and 3' termini of the gene have been determined. Thirty nucleotides before the start of the messenger RNA coding sequence is the heptanucleotide TATATAT, which is also present in a similar location relative to the chicken ovalbumin gene and other unique sequence eucaryotic genes. This sequence resembles that of the Pribnow box in procaryotic genes where a promoter function has been implicated. Seven nucleotides past the 3' end of the gene is the tetranucleotide TTGT, a sequence found to be present at identical locations as either TTTT or TTGT in other eucaryotic genes that have been sequenced. These conserved DNA sequences flanking eucaryotic genes may serve some regulator function in the expression of these genes.     

Nucleotide sequence analysis of the cloned salmon preproinsulin cDNA

A cDNA library was constructed using polyadenylated RNA from salmon (Oncorhynchus keta) Brockmann bodies, plasmid vector pBR322, and in vitro recombinant DNA techniques. Insulin-related clones were identified with a cDNA probe generated from the same RNA and enriched for insulin sequences. Two recombinants were shown to contain the nucleotide sequence of the entire coding region and parts of the 5' and 3' untranslated regions. The salmon preproinsulin mRNA is about 760 nucleotides long, 315 of which code for the protein, while about 190 and 200 nucleotides belong to the 5' and 3' flanking regions, respectively. Comparison of the nucleotide sequences of salmon insulin mRNA with those from other species reveals that sequence conservation is limited to the regions coding for the B and A peptides and two segments of the signal peptide. The C-peptide region exhibits no significant sequence homology with the C-peptides of other vertebrates. The 5' and 3' untranslated regions of the salmon preproinsulin mRNA are homologous only with the anglerfish mRNA, whereas there is no evident homology with those of birds and mammals. In addition to establishing the sequence of the preproinsulin mRNA, cloned salmon insulin cDNA provides a specific probe for the analysis and isolation of genomic DNA fragments containing insulin genes.