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使用基于DNA阵列的比较基因组杂交技术检测单细胞中的非整倍体

Aneuploidy detection in single cells using DNA array-based comparative genomic hybridization.

作者信息

Hu Dong Gui, Webb Graham, Hussey Nicole

机构信息

Reproductive Medicine Unit, Department of Obstetrics and Gynaecology, University of Adelaide, The Queen Elizabeth Hospital, Woodville Road, Woodville, South Australia 5011, Australia.

出版信息

Mol Hum Reprod. 2004 Apr;10(4):283-9. doi: 10.1093/humrep/gah038. Epub 2004 Jan 29.

Abstract

The use of metaphase comparative genomic hybridization (CGH) to screen all human chromosomes for aneuploidy in preimplantation embryos is hindered by the time required to perform the analysis. We report in this paper a novel approach to manufacture a DNA microarray for CGH for the detection of aneuploidy in single cells. We spotted human chromosome-specific libraries on glass slides that were depleted of repetitive sequences and tested our array CGH method in 14 experiments using either single male and/or single female lymphocytes. For the autosomes, the mean normalized ratios were all close to the expected ratio of 1.0 with overall 300/308 (97%) of the normalized ratios falling within the range 0.75 to 1.25. It was possible to deduce the correct copy number of the X chromosome in 13/14 (92.9%) separate array CGH experiments but the Y chromosome in only 4/14 (29%). We tested our microarray CGH method on a single fibroblast from each of three cell lines containing a specific chromosome aneuploidy (trisomy 13, 15 or 18) and in each case our microarray analysis was able to obtain a diagnosis based on the fact that the aneuploid chromosome gave the highest ratio (1.32, 1.27 and 1.27 respectively) with the ratios of all other chromosomes falling within the range 0.75-1.25. Requiring just 30 h, our method may be more suitable for PGD aneuploidy screening than metaphase CGH.

摘要

中期比较基因组杂交(CGH)用于筛查植入前胚胎中所有人类染色体的非整倍体,但该分析所需时间限制了其应用。我们在本文中报告了一种制造用于CGH的DNA微阵列的新方法,用于检测单细胞中的非整倍体。我们将人类染色体特异性文库点样在去除重复序列的载玻片上,并使用单个男性和/或单个女性淋巴细胞在14次实验中测试了我们的阵列CGH方法。对于常染色体,平均标准化比率均接近预期的1.0比率,总体308个标准化比率中有300个(97%)落在0.75至1.25范围内。在14次独立的阵列CGH实验中,有13次(92.9%)能够推断出X染色体的正确拷贝数,但只有4次(29%)能够推断出Y染色体的正确拷贝数。我们在分别含有特定染色体非整倍体(13、15或18三体)的三个细胞系的单个成纤维细胞上测试了我们的微阵列CGH方法,在每种情况下,我们的微阵列分析都能够基于非整倍体染色体给出最高比率(分别为1.32、1.27和1.27)且所有其他染色体的比率落在0.75 - 1.25范围内这一事实做出诊断。我们的方法仅需30小时,可能比中期CGH更适合用于植入前遗传学诊断(PGD)的非整倍体筛查。

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