Nucleotide Sequence of a cDNA Encoding a Heat-Shock Protein (HSP70) from Barley (Hordeum vulgare L.)
Nucleotide Sequence of a cDNA Encoding a Heat-Shock Protein (HSP70) from Barley (Hordeum vulgare L.)
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DOI：
10.1104/pp.106.2.815
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发表时间：
1994-10
期刊：
影响因子：
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通讯作者：
Fuqiang Chen;P. Hayes;D. Mulrooney;A. Pan
中科院分区：
文献类型：
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作者：
Fuqiang Chen;P. Hayes;D. Mulrooney;A. Pan
在各种所谓的热休克蛋白中，有一个高度保守的家族，其平均分子量为70 kD，被称为HSP 70蛋白。一些HSP70是热诱导的，但另一些是组成型的。HSP70已在许多细胞区室中被鉴定，并作为在蛋白质代谢中具有不同作用的分子伴侣发挥作用（综述参见Hendrick和Hartl，1993）。以未授粉和已授粉大麦子房poly（A）+RNA第一链cDNA为探针，构建了大麦授粉子房poly（A）+ RNA cDNA文库。一个2028 bp的cDNA被确定为其授粉后的水平升高，并转化为BluescriptSK体内切除。通过桑格的双脱氧方法测定DNA序列（桑格等人，1977）使用自动测序仪（Applied Biosystems，Foster City，CA，型号373A，版本1.2.0）。序列同源性分析表明该cDNA克隆属于HSP70家族。预测的氨基酸序列与来自菠菜的热休克蛋白的氨基酸序列有87%的同一性（安德森等人，1994年）。大麦HSP 70克隆的特征见表I。在授粉后和胚胎发生早期，观察到mRNA水平升高相比，在未授粉的卵巢。在5个大麦品种（Dicktoo、Morex、Steptoe、哈灵顿和TR 306）的基因组DNA印迹中检测到HSP 7O位点的中度限制性片段长度多态性。使用Steptoe X Morex的大麦基因组作图群体（Kleinhofs等人，1993），将HSP 70基因座定位于大麦6号染色体的正臂，距羧肽酶基因座3.8厘摩（Kleinhofs等，1993年）。
Among various so-called heat-shock proteins is a highly conserved family that has an average molecular mass of 70 kD and is known as the HSP70 proteins. Some HSP70s are heat inducible but others are constitutive. HSP70s have been identified in many cellular compartments and function as molecular chaperones with diverse roles in protein metabolism (for review, see Hendrick and Hartl, 1993). A cDNA library of poly(A)+ RNA from 3-d-old pollinated barley ovaries in Uni-ZAP XR was screened with first-strand cDNA probes of poly(A)+ RNA from unpollinated and pollinated ovaries, respectively. A 2028-bp cDNA was identified for its elevated leve1 after pollination and converted to Bluescript SKby in vivo excision. DNA sequence was determined by the dideoxy method of Sanger (Sanger et al., 1977) using an autosequencer (Applied Biosystems, Foster City, CA, model 373A, version 1.2.0). Sequence homology analysis showed that the cDNA clone belongs to the HSP70 family. The predicted amino acid sequence was 87% identical to that of the heat-shock protein from spinach (Anderson et al., 1994). Characteristics pertaining to the barley HSP70 clone are described in Table I. Elevated mRNA levels were observed after pollination and during early embryogenesis as compared to levels in unpollinated ovaries. Moderate restriction fragment-length polymorphisms were detected for the HSP7O locus in the genomic DNA blots of five barley cultivars (Dicktoo, Morex, Steptoe, Harrington, and TR306). Using the barley genome mapping population of Steptoe X Morex (Kleinhofs et al., 1993), the HSP70 locus was mapped to the plus arm of barley chromosome 6,3.8 centimorgans from the carboxypeptidase locus (Kleinhofs et al., 1993).