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Ananda Anandan Laurence N. Gatehouse Richelle K. Marshall Colleen Murray John T. Christeller 《Plant Molecular Biology Reporter》2009,27(3):355-364
Two variants of the promoter of the squash aspartic acid protease inhibitor multigene family were isolated from Cucurbita maxima cv. ‘Supermarket’ Hybrid genomic DNA. The isolated promoters, possibly not full length, comprised a 5′-untranslated region
(UTR) of 202–208 bp, contained a 63-bp upstream open reading frame (uORF) and the immediate upstream sequences of 441–445 bp.
The two promoters contained several small deletions relative to each other and 22 single base differences but exhibit overall
92.5% homology over 654 bp. When the promoters were fused to a β-glucuronidase reporter gene and expressed in tobacco, one
variant was highly expressed in the companion cells of the inner and outer phloem of leaves and at lower levels in other organs.
The other variant was expressed at high levels in the long glandular trichomes of the leaf. Deletion analysis identified a
region of ~280 bp immediately upstream of the 5′-UTR containing the TATA box that was responsible for phloem specific expression
and a further region of ~180 bp that enhanced expression in one promoter and conferred trichome expression in the other. Removal
of the 5′-UTR, including the uORF, inactivated the phloem promoter. 相似文献
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Qian-Jin Zhou Hong-Li Zhang Xiao-Lei Jiang Ai-Fang Du 《Functional & integrative genomics》2010,10(4):589-601
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Partial amino acid sequences of a 49 kDa apyrase (ATP diphosphohydrolase, EC 3.6.1.5) from the cytoskeletal fraction of etiolated
pea stems were used to derive oligonucleotide DNA primers to generate a cDNA fragment of pea apyrase mRNA by RT-PCR and these
primers were used to screen a pea stem cDNA library. Two almost identical cDNAs differing in just 6 nucleotides within the
coding regions were found, and these cDNA sequences were used to clone genomic fragments by PCR. Two nearly identical gene
fragments containing 8 exons and 7 introns were obtained. One of them (H-type) encoded the mRNA sequence described by Hsieh
et al. (1996) (DDBJ/EMBL/GenBank Z32743), while the other (S-type) differed by the same 6 nucleotides as the mRNAs, suggesting that
these genes may be alleles. The six nucleotide differences between these two alleles were found solely in the first exon,
and these mutation sites had two types of consensus sequences. These mRNAs were found with varying lengths of 3′ untranslated
regions (3′-UTR). There are some similarities between the 3′-UTR of these mRNAs and those of actin and actin binding proteins
in plants. The putative roles of the 3′-UTR and alternative polyadenylation sites are discussed in relation to their possible
role in targeting the mRNAs to different subcellular compartments.
Sequence data from this article were deposited with the DDBJ/EMBL/GenBank Data Libraries under Accession Nos. Genomic sequences
of pea apyrase: AB023621, AB030444, AB030445, AB038554, AB038555. cDNA sequences of pea apyrase: AB022319, AB027614, AB038668,
AB038669. 相似文献
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Yan Shi Xin-Ping Zhu Jing-Kui Yin Qi-Ya Zhang Jian-Fang Gui 《Molecular biology reports》2010,37(3):1483-1493
Interferon-regulatory factor 1 (IRF-1) is the first member of IRF family, which is involved in many biological processes such
as immune response, antiviral defense, cell growth regulation, and apoptosis. In this study, an IRF-1 gene, EcIRF-1, was isolated and characterized from orange-spotted grouper (Epinephelus coioides). The full-length cDNA of EcIRF-1 is 1,730 bp, including an open reading frame of 906 bp, a 5′-terminal untranslated region (5′-UTR) of 153 bp, and a 3′-UTR
of 671 bp. The EcIRF-1 gene consists of 10 exons and 9 introns, spanning over approximate 4.3 kb of genomic sequence. The 5′-UTR sequence contains
an exon and an intron, and the 3′-UTR sequence is included in the last exon. Expression analysis by real-time PCR reveals
that the EcIRF-1 gene is ubiquitously expressed in various healthy fish tissues, whereas its expression is upregulated in vivo in response
to polyinosinic–polycytidylic acid or lipopolysaccharide stimulation. Subcellular localization analysis shows the EcIRF-1 is an intranuclearly localized and immobile protein in the cultured fish cells. Data presented in this paper provide
an important base to further understand EcIRF-1 gene function and its regulation associated with interferon immune system in orange-spotted grouper. 相似文献
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The genomic organization and chromosomal location of theβ-tubulin isogenes inLeishmania donovani promastigotes has been studied by nucleic acid hybridization techniques using a cloned β-tubulin gene. We have cloned aβ-tubulin gene fragment, 3.3 kbp long, from genomic DNA ofLeishmania donovani using a heterologousβ-tubulin DNA as probe. Restriction maps of this clone have been prepared. It has been estimated that there are approximately
11–15 copies of theβ-tubulin genes per haploid genome. The majority of these isogenes are arranged in a tandem repeat with a length of 3.5 kbp
on a single chromosome. In addition a few dispersed gene copies at different chromosomal loci were detected by pulse field
gradient gel electrophoresis. Part of the internal coding region of the gene has been sequenced to confirm the identity of
theβ-tubulin clone and is found to be nearly identical to that ofLeishmania mexicana amazonensis. 相似文献
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Cinnamate 4-hydroxylase (C4H, EC 1.14.13.11) plays an important role in the phenylpropanoid pathway, which produces many economically important secondary
metabolites. A gene coding for C4H, designated as PhC4H (GenBank accession no. DQ211885) was isolated from Parthenocissus henryana. The full-length PhC4H cDNA is 1,747 bp long with a 1,518-bp open reading frame encoding a protein of 505 amino acids, a 40-bp 5′ non-coding region
and a 189-bp 3′-untranslated region. Secondary structure of the deduced PhC4H protein consists of 41.78% alpha helix, 15.64%
extended strand and 42.57% random coil. The genomic DNA of PhC4H is 2,895 bp long and contains two introns; intron I is 205-bp and intron II is 1,172-bp (GenBank accession no. EU440734).
DNA gel blot analysis revealed that there might be a single copy of PhC4H in Parthenocissus henryana genome. By using anchored PCR, a 963-bp promoter sequence was isolated and it contains many responsive elements conserved
in the upstream region of PAL, C4H and 4CL including the P-, A-, L- and H-boxes. 相似文献
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M. E. G. Suykerbuyk P. J. Schaap H. Stam W. Musters J. Visser 《Applied microbiology and biotechnology》1995,43(5):861-870
Rhamnogalacturonase was purified from culture filtrate ofAspergillus aculeatus after growth in medium with sugar-beet pulp as carbon source. Purified protein was used to raise antibodies in mice and with
the antiserum obtained a gene coding for rhamnogalacturonase (rhgA) was isolated from a λ cDNA expression library. The clonedrhgA gene has an open-reading frame of 1320 base pairs encoding a protein of 440 amino acids with a predicted molecular mass of
45 962 Da. The protein contains a potential signal peptidase cleavage site behind Gly-18 and three potential sites forN-glycosylation. Limited homology withA. niger polygalacturonase amino acid sequences is found. A genomic clone ofrhgA was isolated from a recombinant phage λ genomic library. Comparison of the genomic and cDNA sequences revealed that the coding
region of the gene is interrupted by three introns. Furthermore, amino acid sequences of four different peptides, derived
from purifiedA. aculeatus rhamnogalacturonase, were also found in the deduced amino acid sequence ofrhgA.A. aculeatus strains overexpressing rhamnogalacturonase were obtained by cotransformation using either theA. niger pyrA gene or theA. aculeatus pyr A gene as selection marker. For expression of rhamnogalacturonase inA. awamori theA. awamori pyrA gene was used as selection marker. Degradation patterns of modified hairy regions, determined by HPLC, show the recombinant
rhamnogalacturonase to be active, and the enzyme was found to have a positive effect in the apple hot-mash liquefaction process. 相似文献
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The PSI-D subunit is the ferredoxin-binding site of photosystem I, and is encoded by the nuclear genepsaD. We isolated apsaD genomic clone fromNicotiana sylvestris, by screening a genomic library with apsaD cDNA which we previously cloned fromN. sylvestris (Yamamotoet al., Plant Mol Biol 17: 1251, 1991). Nucleotide sequence analysis revealed that this genomic clone contains apsaD gene, which does not correspond to thepsaD cDNA, so we designated these genespsaDb andpsaDa, respectively. ThepsaDb clone encodes a protein of 214 amino acids uninterrupted by introns. The N-terminal sequence determined for theN. sylvestris PSI-D protein encoded bypsaDb begins at the 49th residue. The products ofpsaDa andpsaDb share 82.7% and 79.5% identity at the amino acid and nucleotide levels, respectively. Genomic Southern analysis showed that two copies ofpsaD are present in theN. sylvestris genome. Ribonuclease protection assays and immunoblot analysis inN. sylvestris indicate that both genes are expressed in leaves, stems and flower buds, but neither is expressed in roots. During leaf development, the ratio ofpsaDb topsaDa mRNA increases from 0.12 in leaf buds to 0.36 in mature leaves. The relative abundance of the corresponding proteins decreased over the same developmental period. These results indicate that differential regulation mechanisms controlpsaDa andpsaDb expression at both the mRNA and protein levels during leaf development. 相似文献