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1.
Molecular Cloning and Characterization of the Myostatin Gene in Croceine Croaker, Pseudosciaena crocea 总被引:1,自引:0,他引:1
Myostatin (MSTN) is a negative regulator of skeletal muscle mass and has a potential application in aquaculture. We reported
the characterization of the myostatin gene and its expression in the croceine croaker, Pseudosciaena crocea. The myostatin gene had three exons encoding 376 amino acids. The cDNA was 1,906 bp long with a 5′-UTR and 3′-UTR of 108 bp
and 667 bp, respectively. A microsatellite sequence, CA30 and CA26 separated by TA, existed in the 3′-UTR. Intron I and II were 343 bp and 758 bp in length, respectively. The deduced amino
acid sequence was highly conserved, and had more than 90% identical to shi drum, gilthead seabream, striped sea-bass, white
perch, and white bass proteins. The myostatin of croceine croaker had a putative amino terminal signal sequence (residues
1–22), a transforming growth factor-beta (TGF-β) propeptide domain (residues 41–256), a RXXR proteolytic processing site (RARR,
residues 264–267, matching the RXXR consensus site), and a TGF-β domain (residues 282–376). There were 13 conserved cysteine
residues in croceine croaker myostatin, nine of which are common to all TGF-β superfamily members. The most conserved region
of vertebrate myostatins is the TGF-β domain, which was the mature bioactive domain of the myostatin protein. The myostatin
gene was expressed not only in the skeletal muscle, but also in the other tissues. 相似文献
2.
The TATA-box binding protein (TBP) is one of the 4 DNA-binding proteins that has been shown to associate with the proximal
promoter region (−295) of the gene for bean seed storage protein phaseolin. The −295 promoter is essential for spatial and
temporal control of the phaseolin gene expression. We designed a pair of degenerated primers based on the highly conserved
sequence of the carboxyl-terminal domain of yeast TBP and used PCR to amplify the corresponding sequence from the bean cDNA.
By using the amplified fragment as a probe, we screened a cDNA library derived from poly A(+) RNA from developing bean seeds
and isolated 2 nearly full-length cDNA clones (813 and 826 bp long). The cDNAs encode 2 distinct isoforms of bean TBP, PV1
and PV2, each with an open reading frame of 200 amino acid residues. The 2 cDNA sequences share an 85.8% overall nucleotide
sequence identity, with the coding region showing a higher degree of identity (94.4%) than the 5′- and 3′-untranslated regions
(69%). The deduced amino acid sequence of the bean TBP isoforms differ in only 3 amino acid residues at positions 5, 9, and
16, all located in the amino-terminal region. The carboxyl-terminal domain of 180 amino acid residues shows a high degree
(>82%) of evolutionary sequence conservation with the TBP sequences from other eukaryotic species. This domain possesses the
3 highly conserved structural motifs, namely the 2 direct repeat sequences, a central basic region rich in basic amino acid
residues, and a region similar to the sigma factor of prokaryote. On the basis of this and other findings, we suggest that
higher plants in general may have at least 2 copies of TBP gene, presumably resulting from the global duplication of the genome.
Accession numbers AF015784 and AF015785 at the GenBank. 相似文献
3.
The full sequence of the nitrate reductase gene was obtained from a type isolate of Verticillium fungicola var. fungicola and used for phylogenetic analysis against other ascomycete fungi. Sequencing obtained 2749 bp of coding region, 668 bp of
5′ flanking sequence and 731 bp of 3′ flanking sequence. In silico analysis indicated that the coding region contains a single intron and translates into an 893 amino acid protein, with BLAST
analysis identifying five conserved nitrate reductase domains within the protein. The 5′ flanking sequence contains numerous
conserved sites putatively involved in binding nitrogen regulatory proteins, indicating that the regulation of the gene is
likely to be subject to the same regulation as that of model fungi such as Aspergillus nidulans. The central portion of this gene was amplified and sequenced from a number of V. fungicola isolates and related fungi and the resulting phylogenies compared to those obtained from analysis of the rDNA internal transcribed
spacer regions for these fungi. Both nitrate reductase and ITS analyses provide additional evidence that reinforces previous
findings that suggest the mushroom pathogenic Verticillium species are more related to other chitinolytic fungi such as the insect pathogens Verticillium lecanii and Beauveria bassiana than to the plant pathogenic Verticillia. 相似文献
4.
Liyuan Zhao Tiezhu Mi Yu Zhen Mingyu Li Shanying He Jing Sun Zhigang Yu 《Hydrobiologia》2009,627(1):19-30
5.
Expression Enhancement of a Rice Polyubiquitin Gene Promoter 总被引:11,自引:0,他引:11
An 808 bp promoter from a rice polyubiquitin gene, rubi3, has been isolated. The rubi3 gene contained an open reading frame of 1140 bp encoding a pentameric polyubiquitin arranged as five tandem, head-to-tail
repeats of 76 aa. The 1140 bp 5′ UTR intron of the gene enhanced its promoter activity in transient expression assays by 20-fold.
Translational fusion of the GUS reporter gene to the coding sequence of the ubiquitin monomer enhanced GUS enzyme activity in transient expression assays
by 4.3-fold over the construct containing the original rubi3 promoter (including the 5′ UTR intron) construct. The enhancing effect residing in the ubiquitin monomer coding sequence
has been narrowed down to the first 9 nt coding for the first three amino acid residues of the ubiquitin protein. Mutagenesis
at the third nucleotide of this 9 nt sequence still maintains the enhancing effect, but leads to translation of the native
GUS protein rather than a fusion protein. The resultant 5′ regulatory sequence, consisting of the rubi3 promoter, 5′ UTR exon and intron, and the mutated first 9 nt coding sequence, has an activity nearly 90-fold greater than
the rubi3 promoter only (without the 5′ UTR intron), and 2.2-fold greater than the maize Ubi1 gene promoter (including its 5′ UTR intron). The newly created expression vector is expected to enhance transgene expression
in monocot plants. Considering the high conservation of the polyubiquitin gene structure in higher plants, the observed enhancement
in gene expression may apply to 5′ regulatory sequences of other plant polyubiquitin genes. 相似文献
6.
Cloning, sequencing and expression of cDNA encoding growth hormone from Indian catfish (Heteropneustes fossilis) 总被引:2,自引:0,他引:2
A tissue-specific cDNA library was constructed using polyA+ RNA from pituitary glands of the Indian catfishHeteropneustes fossilis (Bloch) and a cDNA clone encoding growth hormone (GH) was isolated. Using polymerase chain reaction (PCR) primers representing
the conserved regions of fish GH sequences the 3′ region of catfish GH cDNA (540 bp) was cloned by random amplification of
cDNA ends and the clone was used as a probe to isolate recombinant phages carrying the full-length cDNA sequence. The full-length
cDNA clone is 1132 bp in length, coding for an open reading frame (ORF) of 603 bp; the reading frame encodes a putative polypeptide
of 200 amino acids including the signal sequence of 22 amino acids. The 5′ and 3′ untranslated regions of the cDNA are 58
bp and 456 bp long, respectively. The predicted amino acid sequence ofH. fossils GH shared 98% homology with other catfishes. Mature GH protein was efficiently expressed in bacterial and zebrafish systems
using appropriate expression vectors. The successful expression of the cloned GH cDNA of catfish confirms the functional viability
of the clone. 相似文献
7.
Summary The amino acid sequence of the ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO) small subunit (SSU) from Euglena has been established by alignment of the sequence of peptides obtained by cleavage with chymotrypsin, trypsin, Staphylococcus aureus protease or formic acid. The Euglena SSU has 138 amino acids and thus represents longest SSU sequence described so far. Homology is only 41% with cyanobacteria SSU and about 51% with higher plant SSU, whereas it is around 75% between higher plants. The largest homologous portion between all the known SSU sequences is localized in the second half and covers about 20 amino acids. The phylogenetic tree based on known SSU sequences has been established and the rate of amino acid substitution for SSU is estimated to be about 1.35×10-9 per year and per site. Despite heterogeneity in amino acid sequence, we found that the overall secondary structure is fairly well conserved.Abbreviations DABITC
Dimethyl amino azobenzene isothiocyanate
- HPLC
high pressure liquid chromatography
- Kd
Kilo daltons
- LSU
large subunit
- PITC
phenyl isothiocyanate
- RuBisCO
ribulose-1,5-bisphosphate carboxylase/oxygenase
- SDS
sodium dodecyl sulfate
- SSU
small subunit
- TFA
trifluoric acetic acid 相似文献
8.
Yan F Peng J Lu Y Lin L Zheng H Chen H Chen J Adams MJ 《Molecular biology reports》2009,36(6):1283-1289
Dicer-like proteins (DCLs) are involved in small RNA-mediated development and viral defense in plants. In model plants, at
least four DCLs have been found and a number of studies have helped to understand their function. However, the function of
the Dicer or DCLs in other plants is still unclear. Here, we report the full-length cDNA sequence of Brassica rapa ssp. chinensis DCL2 (BrDCL2) gene, which contains a 4,179 bp open reading frame (ORF) encoding a protein of 1,392 amino acids. At the 3′ end of BrDCL2, clones with three different lengths of 3′ untranslated region were found. An alternative splice variant of BrDCL2, BrDCL2sv, in which one intron was retained between exon9 and exon10, was also cloned. Because of a change in the coding sequence resulting
in a premature terminal codon, BrDCL2sv was expected to translate a short peptide containing the whole DEXHc domain. 相似文献
9.
Hua Xie Peirong Xu Yanlong Jia Jie Li Yumin Lu Lexun Xue 《Journal of applied phycology》2007,19(5):497-504
A cDNA corresponding to the nitrate reductase (NR) gene from Dunaliella salina was isolated by RT-PCR and (5′/3′)-RACE techniques. The full-length cDNA sequence of 3,694 bp contained an open reading frame
of 2,703 bp encoding 900 amino acids, a 5′-untranslated region of 151 bp and a 3′-untranslated sequence of 840 bp with a poly
(A) tail. The putative gene product exhibited 78%, 65%, 59% and 50% identity in amino acid sequence to the corresponding genes
of Dunaliella tertiolecta, Volvox carteri, Chlamydomonas reinhardtii, and Chlorella vulgaris, respectively. Phylogenetic analysis showed that D. salina NR clusters together with known NR proteins of the green algae. The molecular mass of the encoded protein was predicted to
be 99.5 kDa, with an isoelectric point of 8.31. This protein shares common structural features with NRs from higher plants
and green algae. The full-length cDNA was heterologously expressed in Escherichia coli as a fusion protein, and accumulated to up to 21% of total bacteria protein. Recombinant NR protein was active in an enzyme
assay, confirming that the cloned gene from D. salina is indeed NR. 相似文献
10.
Cassettes for seed-specific expression tested in transformed embryogenic cultures of soybean 总被引:4,自引:0,他引:4
Soybean (Glycine max [L.] Merrill) lectin is a seed protein that accumulates in protein bodies of cotyledons during seed development. We have
constructed two expression cassettes containing the 5′ and 3′ regions of the soybean lectin gene connected by aNot I restriction site. One vector also contains the 32 amino acid signal sequence. Using polymerase chain reaction (PCR), the
coding region of the β-glucuronidase (uidA) gene was inserted into theNot I site of each vector. We tested the function of the expression cassettes in transformed embryogenic cultures of soybean.
Development-specific GUS expression was observed in developing somatic embryos transformed with the chimeric lectin promoter-GUS
constructs as determined by histochemical assays. Our data indicate that these cassettes could be used to drive expression
of foreign genes to modify embryo-specific traits of soybean as protein quality or quantity in the seed. 相似文献
11.
Y. López H. L. Nadaf O. D. Smith J. P. Connell A. S. Reddy A. K. Fritz 《TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik》2000,101(7):1131-1138
An understanding of the molecular mechanisms that are responsible for increased oleic acid accumulation would open avenues
to alter peanut fatty acid composition and allow detection of polymorphic regions which can be used for marker assisted selection
(MAS). Δ12-Fatty acid desaturase (FAD) was isolated and characterized from genotypes having a low or high oleic to linoleic acid O/L
ratio – genotypes, Tamspan 90 (T-90) and F435–2-2 (F435), respectively. Southern blots showed three to four copies per haploid
genome, and no major differences in organization between the two parental lines. Approximately 3525 bp was isolated from both
genotypes, including a genomic walk toward the promoter region. The Δ12-Fad contains a putative intron, the coding region at the 3′ end, and an open reading frame (ORF) of 1140 bp encoding 379
amino acids. Comparisons of the coding sequences from the high and low oleic acid genotypes revealed several single nucleotide
polymorphisms (SNPs). Two polymorphisms appear to be associated with the high O/L trait. The first is an ”A” insertion 442
bp after the start codon. The ”A” insertion shifts the amino acid reading frame, probably resulting in a truncated, inactive
protein and the loss of one of three histidine boxes believed to be involved in metal ion complexation required for the reduction
of oxygen. Another polymorphism at 448 bp from the start codon results in an amino acid change. The region containing the
polymorphisms was amplified from leaf tissue of several independently derived backcross lines (IDBLs). Most high O/L lines
had either the ”A” insertion or the amino acid substitution.
Received: 1 February 2000 / Accepted: 20 March 2000 相似文献
12.
The techniques of homology cloning and anchored PCR were used to clone the Hsp90 gene from black tiger shrimp. The full length
cDNA of black tiger shrimp Hsp90 (btsHsp90) contained a 5′ untranslated region (UTR) of 72 bp, an ORF (open reading frame)
of 2160 bp encoding a polypeptide of 720 amino acids with an estimated molecular mass of 83-kDa and a 3′ UTR of 288 bp. The
sequence of the coding region showed 90 and 84% homology with that of the Chiromantes haematocheir and Homo sapiens, respectively. Conserved signature sequences of Hsp90 gene family were found in the btsHsp90 deduced amino acid sequence.
The temporal expressions of Hsp90 gene were constitutively in the black tiger shrimp tissues including liver, ovary, muscle,
brain stomach, and heart, and their levels were markedly enhanced after 30-min heat treatment at 37°C. In ovarian maturation
stages, the expression of btsHsp90 was strongest in the second stage, weaker in the fourth and first stage. 相似文献
13.
A gene of a monomeric hemoglobin, the Pol n component of MV, of a chironomid species, Polypedilum nubifer, was cloned by screening
the larval cDNA library with a nucleotide probe corresponding to the N-terminal sequence of purified MV. A clone, 8N, was
755 bp long and comprised a 60 bp 5′ non-coding region, a 209 bp 3′ non-coding region and a 486 bp coding region for 160 amino
acids. A comparison of N-terminal sequence of purified MV with that estimated from the DNA sequence of clone 8N, revealed
the existence of a signal peptide consisting of 14 residues. This signal peptide was almost exclusively composed of hydrophobic
amino acids, suggesting the peptide functions in preglobin transport across the endoplasmic reticulum. The estimated sequence
of mature globin of MV showed only 41% of homology to that of CTT-IV, a chromatographically similar monomeric Hb to MV, of
an another chironomid species, Chironomus thummi thummi, in a 146 alignment. However, displacements in hydrophilic ⇆hydrophobic
manner were observed only at 28 positions whereas those in hydrophobic ⇆hydrophobic or hydrophilic ⇆hydrophilic manner were
observed at 45 positions. Furthermore, a comparison of the haem contact positions between these two Hbs showed a remarkable
conservance and displacements only in hydrophilic ⇆ hydrophilic or hydrophobic ⇆hydrophobic manner, suggesting the crucial
role of these positions in Hb functionality.
This revised version was published online in July 2006 with corrections to the Cover Date. 相似文献
14.
Powdery mildew, caused by Uncinula necator Burr, is one of the most seriously damaging diseases of grapevine all over the world. To gain the novel gene and investigate
the resistance mechanism in Chinese Wild Vitis pseudoreticulata clone Baihe-35-1, mRNA differential display was employed to study the differential expression of the resistant gene to the
disease of it when inoculated by Uncinula necator under natural field conditions, 5′ RACE and 3′ RACE have been used to clone the whole cDNA sequences of VpAPX, the novel
gene related to Ascorbate Peroxidase which involved in resistant to the disease, is composed of specific sequence 1077 bp and has an open reading frame of 750 bp
coding for 250 amino acid residues with a molecular weight of 27.566 kDa. The VpAPX gene was obtained by polymerase chain
reaction (PCR) with the special primers synthesized according to the sequences of cDNA, and further cloned it into the pGEM-T
easy vector. The cloned VpAPX gene was cut out again with two restriction enzymes and was inserted into the prokaryotic expression
vector pGEX-4T-1, then transferred into E. coli BL21. As result, GST-VpAPX fusion protein was successfully expressed by induction of IPTG and purified by GST affinity resin.
After injecting rabbit, the polyclonal antibodies were produced. Western blot analyses showed that the antibody reacted specifically
to GST-VpAPX fusion protein and the titer for this antibody is 105. This research made the foundation to transform the VpAPX gene into grape plants for follow research in processing.
Ling Lin, Xiping Wang: These two authors contributed equally to this work. 相似文献
15.
We have isolated a novel human cDNA coding for human salt-tolerant protein (HSTP), that is a homologue of the rat salt-tolerant
protein (STP) and may contribute to salt-induced hypertension by modulating renal cation transport. The nucleotide sequence
(1988 bp) of the HSTP cDNA contains an open reading frame encoding a polypeptide comprising 545 amino acids, two residues
fewer than the rat STP cDNA. The predicted amino acid sequence exhibits 92% identity to that of the rat protein. HSTP contains
predicted coiled-coil domains and Src Homology 3 domain, and shows a high degree of identity to CIP4 (Cdc42 target protein)
and human Trip 10 (thyroid-hormone receptor interacting protein). We have mapped the HSTPgene to human chromosome 19 by fluorescence in situhybridization.
This revised version was published online in July 2006 with corrections to the Cover Date. 相似文献
16.
The secretogranin II (SCG2) gene is associated with the synthesis and secretion of follicle-stimulating hormone and luteinizing hormone. In the present
study, we have determined the complete cDNA sequence of pig SCG2, which was submitted to GenBank with accession no. AY870646. Its complete open reading frame of 1,851 nucleotides encodes
616 amino acids. The predicted protein shares 80–87% identity with mouse, human, and bovine SCG2 proteins, and all four species share almost complete identity in the secretoneurin and EM66 domains. Pig SCG2 is a protein
of 589 amino acids and 68,132 Da, preceded by a signal peptide of 27 residues. It contains nine pairs of dibasic residues,
which are used as potential cleavage sites for generation of physiologically active peptides. Analysis of the SCG2 gene across the INRA-Minnesota porcine radiation hybrid panel indicates close linkage with microsatellite marker SW2608,
located on Sus scrofa chromosome 15 (SSC15) q25, which harbors several QTL for ovulation rate and meat quality. Comparative sequencing and EST
analysis revealed nine SNPs in porcine SCG2 cDNA, including seven SNPs in the coding region and two SNPs in the 3′ UTR. Four nonsynonymous SNPs (G622A, G1671T, C1718T,
and A1790C) resulted in amino acid substitutions of Ala→Thr, Glu→Asp, Pro→Leu, and Asn→Thr, respectively. 相似文献
17.
R. B. Turley 《Biologia Plantarum》2008,52(4):759-762
Two dimensional polyacrylamide gel electrophoresis (2D-PAGE) was used to identify differentially expressed proteins in wild-type
(DP 5690) and fiberless (SL 1-7-1) cotton ovules. One protein, designated V2 was unique to ovules of the fiber producing DP
5690 line. The protein was purified from 2D-PAGE of 4 d post anthesis DP 5690 ovules and partially sequenced. The short amino
acid sequence was nearly identical to the deduced amino acid sequence for cotton phenylcoumaran benzylic ether reductase (PCBER)
protein. A consensus sequence was assembled from ESTs encoding cotton PCBER genes, primers were designed, and a full length
gene was amplified from plasmid DNA from a 72 h etiolated cotton cotyledon library. The polymerase chain reaction generated
a 950 bp product with unique EcoRI (5′) and (3′) KpnI restriction sites for directional insertion into the expression vector pPICZA. Nucleotide sequencing was performed, and
the full length coding region was 924 bp encoding a protein of 308 amino acids. The molecular mass and pI measured (2D PAGE)
were similar to the theoretical protein. 相似文献
18.
19.
A cDNA encoding a serine proteinase homologue of kuruma shrimp (Marsupenaeus japonicus) was cloned. The 1257 bp cDNA encodes a 339 amino acid putative peptide, with a signal sequence of 16 amino acid residues. The deduced amino acid sequence is 42-67% similar to the immune-related serine proteinases and serine proteinase homologues of arthropods. It contains catalytic triad residues in the putative catalytic domain except for one substitution of Ser by a Gly residue. The six cysteine residues that form three disulphide bridges in most serine proteinases were conserved. The M. japonicus serine proteinase homologue was mainly expressed in haemocytes, in which expression dramatically increased after 3 days feeding with peptidoglycan at 0.2 mg kg(-1) shrimp body weight per day. 相似文献