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Mol. Hum. Reprod. Advance Access published online on October 1, 2004
Molecular Human Reproduction, doi:10.1093/molehr/gah114
© 2004 by Oxford University Press
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Received February 15, 2004
Revised July 13, 2004
Accepted September 12, 2004

Article

Multiple displacement amplification on single cell and possible PGD applications

Ali Hellani 1*, Serdar Coskun 1, Moncef Benkhalifa 2, Abelghani Tbakhi 1, Nadia Sakati 1, Ali Al-Odaib 1, and Pinar Ozand 1

1 King Faisal Specialist Hospital and Research Center, Riyadh, Saudi Arabia
2 ATL R&D Laboratory, Paris, France

* To whom correspondence should be addressed. E-mail: hellani{at}kfshrc.edu.sa.


  Abstract

Multiple displacement amplification (MDA) is a technique used in the amplification of very low amounts of DNA and reported to yield large quantities of high-quality DNA. We used MDA to amplify the whole genome directly from a single cell. The most common techniques used in PGD are PCR and fluorescent in-situ hybridization (FISH). There are many limitations to these techniques including, the number of chromosomes diagnosed for FISH or the quality of DNA issued from a single cell PCR. This report shows, for the first time, use of MDA for single cell whole genome amplification. A total of 16 short tandem repeats (STRs) were amplified successfully with a similar pattern to the genomic DNA. Furthermore, allelic drop out (ADO) derived from MDA was assessed in 40 single cells by analysing (i) heterozygosity for a known {beta} globin mutation (IVSI-5 C-G) and by studying (ii) the heterozygous loci present in the STRs. ADO turned out to be 10.25% for the {beta} globin gene sequencing and 5% for the fluorescent PCR analysis of STRs. Moreover, the amplification accuracy of MDA permitted the detection of trisomy 21 on a single cell using comparative genome hybridization-array. Altogether, these data suggest that MDA can be used for single cell molecular karyotyping and the diagnosis of any single gene disorder in PGD.

Keywords: PGD; whole genome amplification-comparative genome hybridization; CGH-array; multiple displacement amplification.
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