scholarly journals 263 IDENTIFICATION OF A NOVEL MOPT GENE IN HUMAN AND MOUSE ADULT TESTIS

2005 ◽  
Vol 17 (2) ◽  
pp. 281
Author(s):  
Y.-J. Choi ◽  
S.-J. Kang ◽  
J.-H. Kim
Keyword(s):  
Amino Acid ◽  
Genomic Dna ◽  
Wild Type Mouse ◽  
Amino Acid Sequences ◽  
Mouse Tissue ◽  
Pcr Analysis ◽  
Human Testis ◽  
Testicular Development ◽  
Cdna Clones ◽  
Late Spermatid

To discover late stage germ cell-specific transcripts we prepared a cDNA library from adult testes of 35-day old mice and subtracted it with mRNA from the testes of juvenile mice. Real-time RT-PCR analysis indicated that 42 cDNA clones in the subtracted library were expressed more intensely in the adult testes than in the juvenile testes. One clone identified by subtraction is expressed preferentially in the late spermatid and is located on chromosome 17E3 in mouse and 2p22 in human. The full nucleotide and amino acid sequences of mouse and human MOPT gene are deposited in EMBL GenBank (AY367765 And AY367766). Human MOPT is spliced by 5 exons and 4 introns and encompasses 7,000 bp of genomic DNA (from bp 355 822 to 425 511) of NT-022184.13, whereas mouse MOPT is spliced by 5 exons and 4 introns and encompasses 7,382 bp of genomic DNA (from bp 6227407 to 6235588) of NT-039658.2. Because of the limited availability of human testis samples, development-dependent expression of MOPT mRNA was conducted using its mouse homologue and semiquantitative PCR. The number of cycles completed before entering the exponential growth, recorded by amplifier PE5700 for mouse MOPT, were 1.11 ± 0.23, 1.05 ± 0.04, 1.5 ± 0.2, 5.55 ± 0.65, 19.35 ± 0.65, 68.65 ± 2.15, and 185.15 ± 6.15 in W/W, postnatal day 5-, 8-, 12-, 15-, 18-, 22-, and 28-day mouse tissue samples, respectively. The difference among the three times was significant (P < 0.01, ANOVA). These results suggest that expression of MOPT gene increased from postnatal Day 5 to Day 28, indicating possible involvement in testicular development. The ORF encodes a protein containing 79 amino acid residues. A MORN motif, EGQFKDNMFHGLGTYTFPNG, was identified in the predicted protein sequence of MOPT; function of this motif is unknown. In situ hybridization of 12-week-old wild-type mouse testes using an antisense riboprobe and immuno-gold data indicated MOPT was expressed as a late spermatid and acrosome reaction. This work was supported by BK21 program.

scholarly journals Cloning and sequence analysis of cDNAs encoding the α1, α2 and α3 chains of mouse collagen VI

1993 ◽  
Vol 291 (3) ◽  
pp. 787-792 ◽  
Author(s):  
R Z Zhang ◽  
T C Pan ◽  
R Timpl ◽  
M L Chu
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequences ◽  
Untranslated Regions ◽  
Cdna Libraries ◽  
Cdna Clones ◽  
Collagen Vi ◽  
Central Segment ◽  
Glycosylation Sites ◽  
Key Features ◽  
Triple Helical Domain

cDNA clones encoding the alpha 1, alpha 2 and alpha 3 chains of mouse collagen VI have been isolated by screening cDNA libraries with the corresponding human probes. The composite cDNAs for the alpha 1, alpha 2, and alpha 3 chains are 2.5, 1.6 and 2.9 kb in size respectively. The alpha 1 and alpha 2 cDNAs encode the C-terminal portions of the chains as well as the entire 3′-untranslated regions, while the alpha 3 cDNAs encode a central segment of 959 amino acids flanking the triple-helical domain. The deduced amino acid sequences share 86-88% identity with the human counterparts and 67-73% identity with the chicken equivalents. Alignment of the deduced amino acid sequences of mouse, human and chicken collagens reveal that the key features of the protein, including the cysteine residues, imperfections in the Gly-Xaa-Xaa regions, Arg-Gly-Asp sequences and potential N-glycosylation sites, are mostly conserved.

Cloning of rabbit prolactin cDNA and prolactin gene expression in the rabbit mammary gland

1996 ◽  
Vol 16 (1) ◽  
pp. 27-37 ◽  
Author(s):  
L Gabou ◽  
M Boisnard ◽  
I Gourdou ◽  
H Jammes ◽  
J-P Dulor ◽  
...  
Keyword(s):  
Gene Expression ◽  
Mammary Gland ◽  
Amino Acid ◽  
Untranslated Regions ◽  
Pcr Analysis ◽  
Cdna Clones ◽  
Rt Pcr ◽  
Prolactin Gene ◽  
New Zealand White Rabbits ◽  
Rna Probe

ABSTRACT cDNA clones coding for rabbit prolactin were isolated from a pituitary library using a rat prolactin RNA probe. One cDNA contained 873 bases including the entire coding sequence of rabbit prolactin, its signal peptide and the 5′ and 3′ untranslated regions of 44 and 145 nucleotides respectively. The deduced amino acid sequence of the cloned prolactin cDNA presented a 93–78% identity with mink, porcine and human prolactins. The prolactin gene transcription was investigated by RT-PCR analysis in several organs of midlactating New Zealand White rabbits. The ectopic transcription of the prolactin gene was examined in more detail in the mammary gland. A strong PCR signal was detected in the mammary gland of virgin does and was also observed during pregnancy and at the beginning of lactation. This PCR signal was very weak in mid-lactating and absent in post-weaning mammary gland.

scholarly journals Molecular cloning of two human liver 3 α-hydroxysteroid/dihydrodiol dehydrogenase isoenzymes that are identical with chlordecone reductase and bile-acid binder

1994 ◽  
Vol 299 (2) ◽  
pp. 545-552 ◽  
Author(s):  
Y Deyashiki ◽  
A Ogasawara ◽  
T Nakayama ◽  
M Nakanishi ◽  
Y Miyabe ◽  
...  
Keyword(s):  
Bile Acid ◽  
Amino Acid ◽  
Human Liver ◽  
Polyclonal Antibodies ◽  
Amino Acid Sequences ◽  
Cdna Clones ◽  
Amino Acid Residues ◽  
Human Bile ◽  
Coding Regions ◽  
Computer Based

Human liver contains two dihydrodiol dehydrogenases, DD2 and DD4, associated with 3 alpha-hydroxysteroid dehydrogenase activity. We have raised polyclonal antibodies that cross-reacted with the two enzymes and isolated two 1.2 kb cDNA clones (C9 and C11) for the two enzymes from a human liver cDNA library using the antibodies. The clones of C9 and C11 contained coding sequences corresponding to 306 and 321 amino acid residues respectively, but lacked 5′-coding regions around the initiation codon. Sequence analyses of several peptides obtained by enzymic and chemical cleavages of the two purified enzymes verified that the C9 and C11 clones encoded DD2 and DD4 respectively, and further indicated that the sequence of DD2 had at least additional 16 residues upward from the N-terminal sequence deduced from the cDNA. There was 82% amino acid sequence identity between the two enzymes, indicating that the enzymes are genetic isoenzymes. A computer-based comparison of the cDNAs of the isoenzymes with the DNA sequence database revealed that the nucleotide and amino acid sequences of DD2 and DD4 are virtually identical with those of human bile-acid binder and human chlordecone reductase cDNAs respectively.

Expression of intermediate filament proteins during development of Xenopus laevis. I. cDNA clones encoding different forms of vimentin

Development ◽  
1989 ◽  
Vol 105 (2) ◽  
pp. 279-298
Author(s):  
H. Herrmann ◽  
B. Fouquet ◽  
W.W. Franke
Keyword(s):  
Amino Acid ◽  
Xenopus Laevis ◽  
Intermediate Filament ◽  
De Novo ◽  
Mesenchymal Cell ◽  
Amino Acid Sequences ◽  
Cytoskeletal Proteins ◽  
Cdna Clones ◽  
Mammalian Development ◽  
Intermediate Filament Proteins

To provide a basis for studies of the expression of genes encoding the diverse kinds of intermediate-filament (IF) proteins during embryogenesis of Xenopus laevis we have isolated and characterized IF protein cDNA clones. Here we report the identification of two types of Xenopus vimentin, Vim1 and Vim4, with their complete amino acid sequences as deduced from the cloned cDNAs, both of which are expressed during early embryogenesis. In addition, we have obtained two further vimentin cDNAs (Vim2 and 3) which are sequence variants of closely related Vim1. The high evolutionary conservation of the amino acid sequences (Vim1: 458 residues; Mr approximately 52,800; Vim4: 463 residues; Mr approximately 53,500) to avian and mammalian vimentin and, to a lesser degree, to desmin from the same and higher vertebrate species, is emphasized, including conserved oligopeptide motifs in their head domains. Using these cDNAs in RNA blot and ribonuclease protection assays of various embryonic stages, we observed a dramatic increase of vimentin RNA at stage 14, in agreement with immunocytochemical results obtained with antibody VIM-3B4. The significance of very weak mRNA signals detected in earlier stages is discussed in relation to negative immunocytochemical results obtained in these stages. The first appearance of vimentin has been localized to a distinct mesenchymal cell layer underlying the neural plate or tube, respectively. The results are discussed in relation to programs of de novo synthesis of other cytoskeletal proteins in amphibian and mammalian development.

scholarly journals The melanoma antigen gp75 is the human homologue of the mouse b (brown) locus gene product.

1990 ◽  
Vol 171 (4) ◽  
pp. 1375-1380 ◽  
Author(s):  
S Vijayasaradhi ◽  
B Bouchard ◽  
A N Houghton
Keyword(s):  
Amino Acid ◽  
Metastatic Melanoma ◽  
Gene Product ◽  
Coat Color ◽  
Immunological Tolerance ◽  
Amino Acid Sequences ◽  
Cdna Clones ◽  
Igg Antibodies ◽  
Human Homologue ◽  
Melanoma Antigen

The gp75 antigen is an abundant intracellular glycoprotein expressed in melanosomes of human pigmented melanocytes and melanomas. IgG antibodies in sera of a patient with metastatic melanoma have been shown to immunoprecipitate gp75, suggesting that immunological tolerance against gp75 can be broken. The mouse mAb TA99, which specifically recognizes gp75, was used to isolate and purify the antigen. Amino acid sequences of three internal peptides were determined from the purified gp75 polypeptide. cDNA clones were isolated by screening with oligonucleotides based on these peptide sequences. The gp75 peptides and cDNA had approximately 90% identity with, respectively, the derived amino acid and nucleotide sequences of a mouse gene that maps to the b (brown) locus. The brown locus determines coat color in the mouse, suggesting that gp75 regulates or influences the type of melanin synthesized.

Detection of the Na+-translocating NADH-quinone reductase in marine bacteria using a PCR technique

2000 ◽  
Vol 46 (4) ◽  
pp. 325-332 ◽  
Author(s):  
Sanae Kato ◽  
Isao Yumoto
Keyword(s):  
Phylogenetic Analysis ◽  
Amino Acid ◽  
16S Rrna ◽  
Marine Bacteria ◽  
Genomic Dna ◽  
Screening Method ◽  
Quinone Reductase ◽  
Amino Acid Sequences ◽  
Homologous Sequences ◽  
Simple Screening

To examine the distribution of the Na+-translocating NADH-quinone reductase (Na+-NQR) among marine bacteria, we developed a simple screening method for the detection of this enzyme. By reference to the homologous sequences of the Na+-NQR operons from Vibrio alginolyticus and Haemophilus influenzae, a pair of primers was designed for amplification of a part of the sixth ORF (nqr6) of the Na+-NQR operon. When PCR was performed using genomic DNA from 13 marine bacteria, a 0.9-kbp fragment corresponding to nqr6 was amplified in 10 strains. Although there were three PCR-negative strains phylogenetically, based on the sequence of the 16S rRNA, these were placed far from the PCR-positive strains. No product was observed in the case of nonmarine bacteria. The nucleotide and predicted amino acid sequences of nqr6 were highly conserved among the PCR-positive marine bacteria. A phylogenetic analysis of marine bacteria, based on nqr6 sequencing, was performed.Key words: Na+-translocating, NADH-quinone reductase, marine bacteria, PCR.

Serine protease isoforms of Deinagkistrodon acutus venom: cloning, sequencing and phylogenetic analysis

2001 ◽  
Vol 354 (1) ◽  
pp. 161-168 ◽  
Author(s):  
Ying-Ming WANG ◽  
Suei-Rong WANG ◽  
Inn-Ho TSAI
Keyword(s):  
Amino Acid ◽  
Serine Proteases ◽  
Parallel Evolution ◽  
Gel Filtration ◽  
Venom Gland ◽  
Anion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Amino Acid Sequences ◽  
Cdna Clones ◽  
Single Chain

The major coagulating fibrinogenase of Deinagkistrdon acutus venom, designated acutobin, was purified by anion-exchange chromatography, gel filtration and reverse-phase HPLC. Approximately 80% of its protein sequence was determined by sequencing the various fragments derived from CNBr cleavage and digestion with endoprotease. Extensive screening of the venom gland cDNA species after amplification by PCR resulted in the isolation of four distinct cDNA clones encoding acutobin and three other serine proteases, designated Dav-PA, Dav-KN and Dav-X. The complete amino acid sequences of these enzymes were deduced from the cDNA sequences. The amino-acid sequence of acutobin contains a single chain of 236 residues including four potential N-glycosylation sites. The purified acutobin (40kDa) contains approx. 30% carbohydrate by weight, which could be partly removed by N-glycanase. The phylogenetic tree of the complete amino acid sequences of 40 serine proteases from 18 species of Crotalinae shows functional clusters reflecting parallel evolution of the three major venom enzyme subtypes: coagulating enzymes, kininogenases and plasminogen activators. The possible structural elements responsible for the functional specificity of each subtype are discussed.

scholarly journals The two alpha-tubulin genes of Chlamydomonas reinhardi code for slightly different proteins.

1985 ◽  
Vol 5 (9) ◽  
pp. 2389-2398 ◽  
Author(s):  
C D Silflow ◽  
R L Chisholm ◽  
T W Conner ◽  
L P Ranum
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequences ◽  
Cdna Clones ◽  
Gene Products ◽  
Beta Tubulin ◽  
Alpha Tubulin ◽  
Tubulin Genes ◽  
Total Complement ◽  
Cloned Gene ◽  
Net Charges

Full-length cDNA clones corresponding to the transcripts of the two alpha-tubulin genes in Chlamydomonas reinhardi were isolated. DNA sequence analysis of the cDNA clones and cloned gene fragments showed that each gene contains 1,356 base pairs of coding sequence, predicting alpha-tubulin products of 451 amino acids. Of the 27 nucleotide differences between the two genes, only two result in predicted amino acid differences between the two gene products. In the more divergent alpha 2 gene, a leucine replaces an arginine at amino acid 308, and a valine replaces a glycine at amino acid 366. The results predicted that two alpha-tubulin proteins with different net charges are produced as primary gene products. The predicted amino acid sequences are 86 and 70% homologous with alpha-tubulins from rat brain and Schizosaccharomyces pombe, respectively. Each gene had two intervening sequences, located at identical positions. Portions of an intervening sequence highly conserved between the two beta-tubulin genes are also found in the second intervening sequence of each of the alpha genes. These results, together with our earlier report of the beta-tubulin sequences in C. reinhardi, present a picture of the total complement of genetic information for tubulin in this organism.

scholarly journals Molecular cloning of two isoforms of the murine homolog of the myeloid CD33 antigen

Blood ◽  
1994 ◽  
Vol 83 (11) ◽  
pp. 3188-3198 ◽  
Author(s):  
EZ Tchilian ◽  
PC Beverley ◽  
BD Young ◽  
SM Watt
Keyword(s):  
Amino Acid ◽  
Cell Line ◽  
Myeloid Cell ◽  
Amino Acid Identity ◽  
Amino Acid Sequences ◽  
Cdna Clones ◽  
Reading Frame ◽  
Murine Homolog ◽  
Myeloid Cell Line ◽  
Nucleotide Insertion

Abstract CD33 monoclonal antibodies recognize a 67-kD glycoprotein of unknown function that is expressed by early myeloid progenitors and their leukemic counterparts. We report here the cloning of the murine homolog of the human CD33 antigen. Two cDNA clones, differing by an 83- nucleotide insertion in the cytoplasmic region, were isolated. The insertion generated a shift in the reading frame within the cytoplasmic tail, resulting in two mouse CD33 isoforms, m33-A and m33-B, with distinct cytoplasmic domains and with predicted protein core molecular weights of 37 kD and 45 kD, respectively. The cDNAs and deduced amino acid sequences show extensive similarity with the human CD33 sequence with the highest homology occurring in the first and second lg-like domains (61% amino acid identity). The most significant divergence between the human and murine proteins occurs in their cytoplasmic portions. The murine CD33 mRNAs were detected in bone marrow, spleen, thymus, brain, liver, the multipotential progenitor cell line, A4, the myelomonocytic cell line, WEHI3B, the myeloid cell line, M1, and the macrophage cell line, P388, by Northern blot analysis. The expression pattern of the murine CD33 homolog suggests that the function of CD33 antigen in hematopoiesis may be conserved between humans and mice.

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