Hahn S, Garvin A M, Di Naro E, Holzgreve W
Department of Obstetrics and Gynecology, University of Basel, Switzerland.
Genet Test. 1998;2(4):351-5. doi: 10.1089/gte.1998.2.351.
The ability to analyze the genetic material of single cells by the PCR opens up new prospects for diagnostics. Because only two copies of the genetic template are available for amplification, a problem that frequently arises when examining heterozygous loci in single cells is allele drop-out (ADO). ADO results from the preferential amplification of one of a pair of heterozygous alleles, in which the other allele is totally under-represented. In examining single cells from carriers heterozygous for beta-thalassemia mutations, we have found ADO to occur in alleles differing by a single nucleotide, where either the normal or the mutant genotype was absent. We have found that ADO is not overcome by either increasing the amount of DNA template to 20 pg or by primer extension preamplification (PEP), but rather that the best diagnostic accuracy is obtained by examining multiple single cells and basing a diagnosis on the combined results of such an examination.
通过聚合酶链反应（PCR）分析单细胞遗传物质的能力为诊断开辟了新前景。由于用于扩增的遗传模板仅有两份拷贝，在检查单细胞中的杂合位点时经常出现的一个问题是等位基因脱扣（ADO）。ADO是由一对杂合等位基因中一个等位基因的优先扩增导致的，其中另一个等位基因完全未得到充分体现。在检查β地中海贫血突变杂合携带者的单细胞时，我们发现ADO发生在相差单个核苷酸的等位基因中，其中正常或突变基因型缺失。我们发现，将DNA模板量增加到20皮克或通过引物延伸预扩增（PEP）都无法克服ADO，而是通过检查多个单细胞并根据这种检查的综合结果进行诊断可获得最佳诊断准确性。
