Wu S L, Chang T C, Huang C N, Chuang L M, Chang T J
Department of Biochemistry, China Medical College, Taichung, Taiwan.
J Formos Med Assoc. 1998 Sep;97(9):614-8.
To elucidate the germline RET proto-oncogene mutations in Taiwanese families with multiple endocrine neoplasia type 2A (MEN 2A), we extracted DNA from peripheral blood leukocytes of 28 members of two families with MEN 2A. Oligonucleotide primers for exons 10 and 11 were used to analyze the nucleotide sequence of codons 609, 611, 618, and 620 of exon 10, and codon 634 of exon 11 of the RET proto-oncogene. Two fragments of genomic DNA were amplified by polymerase chain reaction (PCR). The amplified PCR products were separated and purified from primers and free nucleotides in agarose gels, and the expected 187-bp and 234-bp bands were cut from the gels and sequenced. Thirteen family members in the two MEN 2A kindreds had mutations in codon 634 of exon 11. In kindred 1 (15 members available for this study), a heterozygous codon 634 mutation in nine members and a homozygous codon 634 mutation in one member led to the substitution of Phe (TTC) for Cys (TGC). Three members of kindred 2 (13 members available for this study) had a heterozygous base pair change in codon 634, which led to the substitution of Arg (CGC) for Cys (TGC). In this study, we found two mutation events occurring in two MEN 2A kindreds and also discovered a homozygous point mutation in one woman that led to heterozygous mutations in all of her children.
为阐明台湾2A型多发性内分泌腺瘤病（MEN 2A）家系中的种系RET原癌基因突变情况，我们从两个MEN 2A家系的28名成员的外周血白细胞中提取了DNA。使用外显子10和11的寡核苷酸引物分析RET原癌基因外显子10的密码子609、611、618和620以及外显子11的密码子634的核苷酸序列。通过聚合酶链反应（PCR）扩增基因组DNA的两个片段。将扩增的PCR产物在琼脂糖凝胶中与引物和游离核苷酸分离并纯化，从凝胶中切下预期的187 bp和234 bp条带并进行测序。两个MEN 2A家系中的13名家庭成员在外显子11的密码子634处发生了突变。在家族1（本研究中有15名成员可供分析）中，9名成员存在杂合子密码子634突变，1名成员存在纯合子密码子634突变，导致苯丙氨酸（TTC）取代半胱氨酸（TGC）。家族2（本研究中有13名成员可供分析）的3名成员在密码子634处发生了杂合碱基对变化，导致精氨酸（CGC）取代半胱氨酸（TGC）。在本研究中，我们在两个MEN 2A家系中发现了两个突变事件，还发现一名女性存在纯合子点突变，导致其所有子女均出现杂合子突变。
