Stackhouse R, Chandra T, Robson K J, Woo S L
J Biol Chem. 1983 Jan 25;258(2):703-6.
Antithrombin III mRNA was enriched from a baboon liver by specific polysome immunoprecipitation. The partially purified antithrombin III mRNA preparation was used for cDNA synthesis and cloning. Candidate antithrombin III cDNA clones were identified by differential hybridization using as probes [32P]cDNAs synthesized from the polysome-enriched and -depleted RNA fractions, respectively. The candidate clones were further analyzed by hybrid-selected translation. The authenticity of a cDNA clone positive to both tests was unambiguously confirmed by matching its nucleotide sequence with the known amino acid sequence of human antithrombin III. The baboon antithrombin III cDNA clone hybridized well with human antithrombin III mRNA and can be used as a probe to isolate the corresponding human gene.
通过特异性多核糖体免疫沉淀从狒狒肝脏中富集抗凝血酶III mRNA。将部分纯化的抗凝血酶III mRNA制剂用于cDNA合成和克隆。分别使用从多核糖体富集和耗尽的RNA组分合成的[32P]cDNA作为探针，通过差异杂交鉴定候选抗凝血酶III cDNA克隆。通过杂交选择翻译进一步分析候选克隆。通过将其核苷酸序列与已知的人抗凝血酶III氨基酸序列进行匹配，明确证实了对两种测试均呈阳性的cDNA克隆的真实性。狒狒抗凝血酶III cDNA克隆与人抗凝血酶III mRNA杂交良好，可作为探针分离相应的人类基因。
