Schloesser M, Slomski R, Wagner M, Reiss J, Berg L P, Kakkar V V, Cooper D N
Institut für Humangenetik, Universität Göttingen, F.R.G.
Mol Biol Med. 1990 Dec;7(6):519-23.
Both normal and pathological transcripts of tissue-specific genes may be detected by polymerase chain reaction (PCR) amplification in tissues not normally considered to express the gene product. The exploitation of constitutive basal mRNA levels ("ectopic" transcription) would be a major boon to diagnostic medicine since it promises both to simplify the analysis of complex genes and to avoid the requirement for an expressing tissue that is sometimes obtainable only by biopsy. We have demonstrated the feasibility of this novel strategy by characterizing a mutation in the X-chromosomal Duchenne (or Becker) muscular dystrophy (DMD/BMD) gene encoding dystrophin. The massive size of this gene has in the past often hindered carrier detection due to the high frequency of recombination and the high proportion of new mutations. In this study a deletion was identified in both a BMD patient and a heterozygous carrier using only a minimal volume of peripheral blood. Following specifically primed reverse transcription of lymphocyte RNA, the relevant region of the pathological cDNA was PCR-amplified. Sequence analysis indicated an in-frame deletion of exons 45 to 47.
通过聚合酶链反应（PCR）扩增，在通常不被认为表达该基因产物的组织中，可能检测到组织特异性基因的正常和病理转录本。利用组成型基础mRNA水平（“异位”转录）对诊断医学将是一大福音，因为它有望简化复杂基因的分析，并避免对有时只能通过活检获取的表达组织的需求。我们通过鉴定编码抗肌萎缩蛋白的X染色体杜兴氏（或贝克氏）肌营养不良症（DMD/BMD）基因中的一个突变，证明了这种新策略的可行性。由于重组频率高和新突变比例高，该基因的巨大规模过去常常阻碍携带者检测。在本研究中，仅使用少量外周血，就在一名BMD患者和一名杂合子携带者中鉴定出了一个缺失。在淋巴细胞RNA经特异性引物逆转录后，对病理cDNA的相关区域进行PCR扩增。序列分析表明外显子45至47发生了框内缺失。
