Vuorio E, Sandell L, Kravis D, Sheffield V C, Vuorio T, Dorfman A, Upholt W B
Nucleic Acids Res. 1982 Feb 25;10(4):1175-92. doi: 10.1093/nar/10.4.1175.
Type II procollagen mRNA has been partially purified from embryonic chick sternal cartilage by guanidine hydrochloride extraction, sucrose gradient sedimentation and Sepharose 4B chromatography. Double stranded cDNA was synthesized using AMV reverse transcriptase and E. coli DNA polymerase I, tailed using terminal transferase and inserted into the Pst I site of pBR322. Two putative type II procollagen cDNA clones have been characterized. Both plasmids hybridize to 2 sternal RNA species, a major species of 5.3 kb and a minor species of 7 kb. These RNAs are present in total RNA from sterna and differentiated limb bud cultures but are not detected in RNA from stage 24 limb bud which has not yet differentiated to cartilage or in RNA from calvaria. The time of appearance of these RNAs during the differentiation of limb mesenchyme in culture parallels the appearance of translatable type II procollagen mRNA.
Ⅱ型前胶原信使核糖核酸已通过盐酸胍提取、蔗糖梯度沉降和琼脂糖4B层析从胚胎鸡胸骨软骨中部分纯化出来。使用禽成髓细胞瘤病毒逆转录酶和大肠杆菌DNA聚合酶I合成双链互补脱氧核糖核酸，用末端转移酶加尾并插入pBR322的Pst I位点。已鉴定出两个假定的Ⅱ型前胶原互补脱氧核糖核酸克隆。两种质粒都与两种胸骨核糖核酸种类杂交，一种主要的5.3千碱基种类和一种次要的7千碱基种类。这些核糖核酸存在于胸骨和分化的肢芽培养物的总核糖核酸中，但在尚未分化为软骨的24期肢芽的核糖核酸或颅骨的核糖核酸中未检测到。在培养的肢体间充质分化过程中这些核糖核酸出现的时间与可翻译的Ⅱ型前胶原信使核糖核酸的出现平行。
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