A KpnI DNA polymorphism in the human von Willebrand factor (VWF) gene.
A KpnI DNA polymorphism in the human von Willebrand factor (VWF) gene.
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人类血管性血友病因子 (VWF) 基因中的 KpnI DNA 多态性。
DOI：
10.1093/nar/18.16.4968-a
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发表时间：
1990
影响因子：
14.9
通讯作者：
Hilgartner,MW
中科院分区：
文献类型：
--
作者：
Driscoll,CM;Chiu,C;Hilgartner,MW
描述、来源和方法：迄今为止，仅报道了 I 型原胶原 (COLlAI) 的 proa I (I) 链基因中的一个 RFLP (1)。使用跨越 proc 1 (I) C-前肽中的 EcoRI 位点的 5' 引物 (5'-AGACCAGGAATTCGGCTTCG-3') 和针对核苷酸 + 203 至 + 222 的 3' 引物 (5'-TTGGATCCAAGGTTGAATGCACTTTTGG-3') 对 cDNA 和基因组 DNA 进行 PCR 扩增（+ 1 用于标注第一个） 终止密码子后的核苷酸）并含有额外的 BamHI 位点。使用商业试剂盒(Cetus/Perkin-Elmer)和DNA热循环仪在94℃(1分钟)、52℃(1分钟)和72℃(1分钟)下进行25个扩增循环。用EcoRI和BamHI消化后，将265个碱基对片段克隆到M13中并测序。通过等位基因特异性寡核苷酸杂交来区分这两个等位基因 (2)。对带有C的等位基因特异的寡核苷酸具有序列5'-TGAACCCCCCCAAAAGCCA-3'，对带有T的等位基因特异的寡核苷酸具有序列5'-TGGCTTTTGAGGGGGTCA-3'。多态性：序列多态性位于前(I)链mRNA的+88位(+1用于标记终止密码子后的第一个核苷酸)。 + 88 处的核苷酸是 C 或 T（图 1）。 频率：研究无关个体：研究了来自一个家族的 7 个个体、另一个家族的 3 个个体和 8 个无关个体的染色体。其中 5 个个体是任一等位基因的杂合子，11 个个体是 T 纯合子，2 个个体是 + 88 位的 C 纯合子。
Description, Source and Method: To date, only one RFLP (1) has been reported in the gene for the proa I (I) chain of type I procollagen (COLlAI). cDNA and genomic DNA was PCR amplified with a 5'primer (5'-AGACCAGGAATTCGGCTTCG-3') spanning the EcoRI site in the proc 1 (I) C-propeptide and a 3'primer (5'-TTGGATCCAAGGTTGAATGCACTTTTGG-3') directed to nucleotides+ 203 to+ 222 (+ 1 being used to note the first nucleotide after the stop codon) and containing an additional BamHI site. Twenty-five cycles of amplification were performed at 94 C (1 min), 52 C (1 min), and 72 C (1 min) using a commercial kit (Cetus/Perkin-Elmer), and a DNA thermocycler. After digestion with EcoRl and BamHI, the 265 base pair fragment was cloned into M13 and sequenced. The two alleles were distinguished by allele specific oligonucleotide hybridization (2). The oligonucleotide specific for the allele with a C had the sequence 5'-TGAACCCCCCCAAAAGCCA-3'and the oligonucleotide specific for the allele with a T had the sequence 5'-TGGCTTTTGAGGGGGTTCA-3'. Polymorphism: The sequence polymorphism was at position+ 88 (+ 1 being used to note the first nucleotide after the stop codon) of the proal (I) chain mRNA. The nucleotide at+ 88 was either a C or a T (Fig. 1).Frequency: Unrelated individuals studied: chromosomes from seven individuals ofone family, from three individuals of another family, and from eight unrelated individuals were studied. Five individuals were heterozygotes for either allele, eleven were homozygote for a T and two homozygotes for a C at position+ 88.