Isolation and characterization of the mouse metallothionein-I gene.
Isolation and characterization of the mouse metallothionein-I gene.
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小鼠金属硫蛋白-I 基因的分离和表征。
DOI：
10.1073/pnas.77.11.6511
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发表时间：
1980
影响因子：
11.1
通讯作者：
Palmiter,RD
中科院分区：
文献类型：
--
作者：
Durnam,DM;Perrin,F;Gannon,F;Palmiter,RD
双链 cDNA 从富含金属硫蛋白 mRNA 活性的小鼠肝脏 mRNA 级分中合成，连接至限制性位点接头，插入 pBR322，并用于转化大肠杆菌 chi 1776。确定了含有与金属硫蛋白 mRNA 杂交的 DNA 的最大质粒的序列，并显示其含有 380 个碱基对的插入片段，其中包括整个编码区和 3' 非翻译区金属硫蛋白-I。金属硫蛋白-I 插入物经过切口翻译，用于筛选 lambda 噬菌体中的小鼠骨髓瘤和小鼠胚胎 DNA 文库。从每个文库中分离出含有 13-15 kb 小鼠 DNA 的金属硫蛋白-I 基因组克隆。两者均含有 3.8 千碱基对的 EcoRI 片段，可与金属硫蛋白-I 探针杂交。通过异源双链体和限制性图谱确定 3.8 千碱基对片段内金属硫蛋白-I 基因的位置、大小和方向。该基因跨越 1.1 千碱基对并包含至少两个内含子。
Double-stranded cDNA was synthesized from a mouse liver mRNA fraction enriched for metallothionein mRNA activity, ligated to restriction site linkers, inserted into pBR322, and used to transform Escherichia coli chi 1776. The sequence of the largest plasmid containing DNA that hybridized to metallothionein mRNA was determined and shown to contain a 380-base-pair insert that includes the entire coding region and 3' untranslated region of metallothionein-I. The metallothionein-I insert was nick-translated and used to screen both a mouse myeloma and a mouse embryo DNA library in bacteriophage lambda. A metallothionein-I genomic clone containing 13-15 kilobase pairs of mouse DNA was isolated from each library. Both contain a 3.8-kilobase-pair EcoRI fragment that hybridizes to the metallothionein-I probe. The location, size, and orientation of the metallothionein-I gene within the 3.8-kilobase-pair fragment were determined by heteroduplex and restriction mapping. The gene spans 1.1 kilobase pairs and contains at least two introns.