- 期刊
Detection of Dystrophin Gene Deletion in Chinese Duchenne/Becker Muscular Dystrophy Patients Utilizing Multiplex Polymerase Chain Reaction
利用基因增幅技術偵測杜顯型及貝克型營養性肌腇縮症病人之基因缺失
摘要
杜顯型肌肉症（DMD）及貝克型肌肉萎縮症（DMD）是屬於性連隱性遺傳的先天性肌肉疾患。近年來，Kunkel等及Worton等人已將其遺傳基因所在研究成功。利用幾種cDNAprobe做RFLP分析，能診斷出帶變異基因者，或做出生前的診斷。1995年，一種利用基因增幅技術的polymerase chain reaction（PCR）法被公諸於世，利用此法能將基因缺損的有無，快速的、正確的且不用放射性質的加以診斷。在此研究中，吾等利用此法對上述疾病做基因缺失的診斷研究。病人對象為本學院附設醫院小兒科、神經科診斷為DMD或BMD共41位病人，並以8位正常人為對照組。經周邊血管抽取靜脈10ml，利用phenol/chloroform的方法抽取並純化genomic DNA。Oligonucleotide primers利用DNA合成儀為之。複合PCR法是利用Sakuraba等人的方法稍作修改，即將11組primer分為二群，分別進行之。結果顯示，於正常人取得的genomicDNA中，經PCR法增幅後可得到與預其分子量相同的11條DNA產物。而在在病患組中，在排除偽陰性的可能性後，有15位（37%）證實有不同程度的缺失。其中1位缺失在Exon 6和8，1位失在Exon 8，1位缺失在Exon 8，13和17，1位缺失在Exon 17，3位缺失在Exon 43，1位缺失在Exon 50，3位缺失在Exon 50和52。DMD和BMD病人無論在缺失位置或缺失大小，均沒有明顯的差異。
關鍵字
無資料並列摘要
Multiplex PCR was ultilized to detect deletions at the hot spot regions of dystrophin gene. Thirty-four cases of DMD and 7 cases of BMD, based on their clinical manifestations, were examined. Eight normal male adults were included as control. Eleven DNA fragments coding the exons 1, 3, 6, 8, 13, 17, 43, 47, 50, 52 and 60 were chosen and primers were prepared with oligonucleotide synthesizer. Multiplex PCR was performed under the condition of 94¢XC 1 minute for denaturation, 65¢XC 3 minutes for annealing, 72¢XC 3 minutes for extension. Thirty cycles were followed by a last cycle with 7 minutes extension. Clinically, there were no significant differences between Chinese patients and those reported in other areas except the low rate of positive family history (32%). The results from multiplex PCR showed that 15 cases (37%), 11 DMD and 4 BMD, proved to have deletions in the exons studied; 1 located at exons 6 and 8, 1 at exon 8, 1 at exons 8, 13 and 17, 1 at exons 13 and 17, 1 at exon 17, 3 at exon 43, 1 at exon 50, and 3 at exons 50 and 52, after false negtives were excluded. No difference in size or location was noticed between DMD and BMD in the sample-limited result.