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首页> 外文学位 >Characterization of INF56, a gene expressed during infection structure development of the bean rust fungus, Uromyces appendiculatus.
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Characterization of INF56, a gene expressed during infection structure development of the bean rust fungus, Uromyces appendiculatus.

机译：INF56的特征，INF56是在豆类锈菌乌头霉菌感染结构发育过程中表达的基因。

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摘要

Expression of the INF56 gene is induced after the initiation of appressorium formation in bean rust germlings, Uromyces appendiculatus. The objectives of the research are to characterize the gene, and to investigate its possible function in infection structure differentiation.; INF 56 was first subcloned as a 2.6kb fragment and sequenced. The location of the 1.0kb transcript was determined by S1 nuclease protection and primer extension. A TATA box is located 38bp upstream and a CAAT box 130bp upstream of the major transcription initiation site. The gene contains two open reading frames, ORF1 and ORF2. ORF1 is predicted to encode a 14.1kD polypeptide. The amino acid sequence is rich in Ser, Gly and Pro, and it has certain sequence similarity to a functional domain of mammalian cytokeratin type II. ORF2 encodes a 10.1kD polypeptide, is rich in Pro, and it has certain sequence similarity to the cell surface recognition region of chicken fibronectin. ORF1 is more biased than ORF2 for pyrimidine in the third position of each codon. The two open reading frames share one 67bp intron. As identified by hybrid-selection and in vitro translation system, the gene codes for two polypeptides with apparent molecular weight of 15.5kD and 23kD on SDS-PAGE. The gene does not share homology with other plant pathogenic Ascomycetes and Basidiomycetes except for cowpea rust, Uromyces vignae, a subspecies of bean rust.; Sequence comparison between INF56 genomic DNA and cDNA revealed some base changes (1%) randomly spread throughout the coding region of the gene. Southern analyses of genomic DNA probed with the cDNA showed that the rust genome contains at least four copies of the gene. Since the uredospore is dikaryotic, this indicated that there may be four alleles in the bean rust genome, two alleles located on each of two loci. Further evidence suggested that the INF56 gene family is relatively homologous with a few restriction site differences. All copies contain the intron.; The INF56 cDNA was fused with TrpE promoter in a pATH expression vector. Two fusion proteins derived from ORF1 and ORF2 of INF56 were expressed in E. coli, isolated and purified individually for induction of specific antibodies. The antibody will be characterized using Western blots and will be used for cytological studies in the future.
机译：在豆锈菌的幼虫，Uromyces appendiculatus中，发生了内生器官的形成，诱导了INF56基因的表达。该研究的目的是表征该基因，并研究其在感染结构分化中的可能功能。首先将INF 56亚克隆为2.6kb片段并测序。通过S1核酸酶保护和引物延伸确定1.0kb转录物的位置。 TATA盒位于主要转录起始位点上游38bp，CAAT盒位于130bp上游。该基因包含两个开放阅读框，ORF1和ORF2。预测ORF1编码14.1kD多肽。该氨基酸序列富含Ser，Gly和Pro，并且与II型哺乳动物细胞角蛋白的功能域具有一定的序列相似性。 ORF2编码10.1kD多肽，富含Pro，并且与鸡纤连蛋白的细胞表面识别区域具有一定的序列相似性。在每个密码子的第三个位置，嘧啶的ORF1比ORF2更偏。两个开放阅读框共享一个67bp的内含子。如通过杂交选择和体外翻译系统所鉴定，该基因编码在SDS-PAGE上具有表观分子量为15.5kD和23kD的两种多肽。该基因与其他植物病原体子囊菌和担子菌没有同源性，除了cow豆锈，豆腐亚种Uromyces vignae。 INF56基因组DNA和cDNA的序列比较显示一些碱基变化（1％）随机分布在基因的整个编码区域。用该cDNA探测的基因组DNA的Southern分析表明，锈基因组至少包含该基因的四个拷贝。由于脲孢子是双核的，这表明在豆锈基因组中可能存在四个等位基因，两个等位基因位于两个基因座的每个基因座上。进一步的证据表明，INF56基因家族相对同源，具有一些限制性位点差异。所有副本均包含内含子。在pATH表达载体中将INF56 cDNA与TrpE启动子融合。两种衍生自INF56的ORF1和ORF2的融合蛋白在大肠杆菌中表达，分别进行分离和纯化以诱导特异性抗体。该抗体将使用Western印迹进行表征，并将在将来用于细胞学研究。

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  • 作者

    Xuei, Xiaoling.;

  • 作者单位

    Cornell University.;

  • 授予单位 Cornell University.;
  • 学科 Agriculture Plant Pathology.; Biology Molecular.; Biology Genetics.
  • 学位 Ph.D.
  • 年度 1991
  • 页码 153 p.
  • 总页数 153
  • 原文格式 PDF
  • 正文语种 eng
  • 中图分类 植物病理学;分子遗传学;遗传学;
  • 关键词

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