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首页> 外文期刊>Proceedings of the National Academy of Sciences of the United States of America >Identification of cDNA clones encoding valosin-containing protein and other plant plasma membrane-associated proteins by a general immunoscreening strategy.
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Identification of cDNA clones encoding valosin-containing protein and other plant plasma membrane-associated proteins by a general immunoscreening strategy.

机译：通过常规免疫筛选策略鉴定编码含缬沙蛋白的蛋白和其他植物质膜相关蛋白的cDNA克隆。

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摘要

An approach was developed for the isolation and characterization of soybean plasma membrane-associated proteins by immunoscreening of a cDNA expression library. An antiserum was raised against purified plasma membrane vesicles. In a differential screening of approximately 500,000 plaque-forming units with the anti-(plasma membrane) serum and DNA probes derived from highly abundant clones isolated in a preliminary screening, 261 clones were selected from approximately 1,200 antiserum-positive plaques. These clones were classified into 40 groups by hybridization analysis and 5'- and 3'-terminal sequencing. By searching nucleic acid and protein sequence data bases, 11 groups of cDNAs were identified, among which valosin-containing protein (VCP), clathrin heavy chain, phospholipase C, and S-adenosylmethionine:delta 24-sterol-C-methyltransferase have not to date been cloned from plants. The remaining 29 groups did not match any current data base entries and may, therefore, represent additional or yet uncharacterized genes. A full-length cDNA encoding the soybean VCP was sequenced. The high level of amino acid identity with vertebrate VCP and yeast CDC48 protein indicates that the soybean protein is a plant homolog of vertebrate VCP and yeast CDC48 protein.
机译：开发了一种通过cDNA表达文库的免疫筛选来分离和表征大豆质膜相关蛋白的方法。产生针对纯化的质膜囊泡的抗血清。用抗（质膜）血清和DNA探针（初步筛选中分离出的高度丰富的克隆）衍生的约500,000个噬菌斑形成单位的差异筛选中，从约1,200个抗血清阳性噬菌斑中选择了261个克隆。通过杂交分析和5'-和3'-末端测序将这些克隆分为40组。通过搜索核酸和蛋白质序列数据库，鉴定出11组cDNA，其中含valosin的蛋白（VCP），网格蛋白重链，磷脂酶C和S-腺苷甲硫氨酸：δ24-甾醇-C-甲基转移酶不需要日期已从植物中克隆。其余29个组与任何当前数据库条目均不匹配，因此可能代表其他或尚未表征的基因。对编码大豆VCP的全长cDNA进行测序。与脊椎动物VCP和酵母CDC48蛋白的氨基酸高度同一性表明，大豆蛋白是脊椎动物VCP和酵母CDC48蛋白的植物同源物。

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