Mutation analysis of the entire replicated portion of PKD1 using genomic DNA samples
Mutation analysis of the entire replicated portion of PKD1 using genomic DNA samples
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DOI：
10.1681/asn.v125955
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发表时间：
2001-05-01
影响因子：
13.6
通讯作者：
Germino, GG
中科院分区：
文献类型：
--
作者：
Phakdeekitcharoen, B;Watnick, TJ;Germino, GG
PKD1的复制部分占基因长度的近70%，预计在受影响的常染色体显性多囊肾病1型家系中至少有85%的突变存在于PKD1的复制部分。报道的涉及该片段的突变相对较少是由于该基因的基因组结构带来的重大技术挑战。以前基于基因组dna的策略无法评估外显子1和22，并且依赖于使用10至13 kb的PCR产物。在本报告中，描述了一组六种新的引物对组合，它们可以与先前报道的试剂一起用于分析复制区域（外显子1至34）的所有外显子。没有任何产物的长度大于5.8 kb，并且可以使用各种引物组合将该长度减少一半。利用这种方法，鉴定出两种新的致病突变，四种新的疾病相关错义替换和六种新的正常变异。这些新试剂应该对有兴趣对这种疾病进行DNA测试的研究人员有用。
The replicated portion of PKD1, which comprises nearly 70% of the length of the gene, is predicted to harbor at least 85% of the mutations present in affected autosomal dominant polycystic kidney disease type 1 pedigrees. The relative paucity of reported mutations involving this segment is attributable to the significant technical challenges posed by the genomic structure of the gene. Previous genomic DNA-based strategies were unable to evaluate exons 1 and 22 and relied on the use of 10- to 13-kb PCR products. In this report, a set of six novel primer pair combinations, which can be used with previously reported reagents to analyze all of the exons in the replicated region (exons 1 to 34), are described. No product is greater than 5.8 kb in length, and various primer combinations can be used to reduce this length in half. Using this approach, two new pathogenic mutations, four novel disease-associated missense substitutions, and six new normal variants were identified. These new reagents should prove useful to investigators interested in performing DNA testing for this disorder.