Carter N P, Fiegler H, Piper J
Wellcome Trust Sanger Institute, Hinxton, United Kingdom.
Cytometry. 2002 Oct 1;49(2):43-8. doi: 10.1002/cyto.10153.
Array-comparative genomic hybridization (CGH), although providing much higher resolution compared with conventional CGH, has not yet become a widely applied method for the analysis of genomic gains and losses.
In January 2002, the Wellcome Trust sponsored a workshop where many of the laboratories developing this technology met to compare different methodologies for array-CGH. Fourteen groups participated, comprising 11 from Europe and 3 from the United States. To facilitate objective analysis, each laboratory constructed arrays using the same anonymous clones and performed a series of test hybridizations using identical genomic DNAs.
A figure of merit (FM) was developed to summarize entire collections of data from each laboratory in a single measurement. The FMs consistently showed that a few groups produced quantitative array hybridization data of high quality, whereas a majority achieved a lower standard.
The conclusions of the workshop were that polymerase chain reaction-based methods for the amplification of large insert clones for arraying were effective for array-CGH. It was also concluded that hybridizations performed under coverslips or in automated hybridization apparatus were less effective than hybridizations performed in simple wells with gentle rocking. A common experience by the participants was the batch-to-batch variability of commercial Cot1 preparations in their ability to suppress hybridization to repeat sequences. (Supplementary material for this article can be found in the online issue, which is available at http://www.interscience.wiley.com/jpages/0196-4763/suppmat/49_2/v49.43.html or at http://www.sanger.ac.uk/HGP/Cytogenetics/Publications/Cytometry Sept 2002/Supplemental.pdf.)
与传统比较基因组杂交(CGH)相比,阵列比较基因组杂交(array-CGH)虽然分辨率高得多,但尚未成为分析基因组增减的广泛应用方法。
2002年1月,威康信托基金会主办了一次研讨会,许多开发这项技术的实验室齐聚一堂,比较阵列CGH的不同方法。14个小组参与其中,包括11个来自欧洲的小组和3个来自美国的小组。为便于客观分析,每个实验室使用相同的匿名克隆构建阵列,并使用相同的基因组DNA进行一系列测试杂交。
制定了一个品质因数(FM),以单一测量总结每个实验室的全部数据。品质因数始终表明,少数小组产生了高质量的定量阵列杂交数据,而大多数小组达到的标准较低。
研讨会的结论是,基于聚合酶链反应的方法用于扩增用于阵列的大插入片段克隆对阵列CGH是有效的。还得出结论,在盖玻片下或自动杂交仪器中进行的杂交不如在简单孔中轻轻摇动进行的杂交有效。参与者的一个共同经验是,商业Cot1制剂在抑制与重复序列杂交的能力方面存在批次间差异。(本文的补充材料可在在线期刊中找到,网址为http://www.interscience.wiley.com/jpages/0196-4763/suppmat/49_2/v49.43.html或http://www.sanger.ac.uk/HGP/Cytogenetics/Publications/Cytometry Sept 2002/Supplemental.pdf。)