You Position: Home > Paper

Influence of different amplification methods and probes with various lengths on the results of comparative genomic hybridization analysis of preimplanted single blastomere

( views:378, downloads:100 )
Author:
No author available
Journal Title:
CHINESE JOURNAL OF PERINATAL MEDICINE
Issue:
5
DOI:
10.3760/cma.j.issn.1007-9408.2011.05.005
Key Word:
分裂球;核酸杂交;核酸扩增技术;核酸探针;聚合酶链反应;植入前诊断;Blastomeres;Nucleic acid hybridization;Nucleic acid amplification techniques;Nucleic acid probes;Polymerase chain reaction;Preimplantation diagnosis

Abstract: Objective To investigate the effect of different amplification methods and probes with various length on the results of comparative genomic hybridization (CGH) analysis of pre-implanted single blastomere and to establish the basis for preimplantation genetic diagnosis.Methods Twenty blastomeres of embryo at 6-8 cells stage were randomly divided into A and B group with 10 in each.Twenty peripheral blood lymphocytes from a healthy man were similarly divided into C and D group with 10 in each.Degenerate oligonucleotide primed polymerase chain reaction (DOP-PCR) was used to amplify whole genomic DNA in group A and C,and multiple displacement amplification (MDA) was used in group B and D for whole genome amplification (WGA).The specificity of resultant products was confirmed by amplification of TBX1 gene exon 2.CGH was performed respectively with 250-750 bp and 750-2000 bp probes prepared from the amplified whole genomic DNA.The result of CGH was verified by sex-determining region of Y (SRY).Results (1) Nine of the 10 samples in group A and all in group C were amplifiable by DOP-PCR,but there were multiple non-specific bands in the amplification of TBX1 exon 2 when WGA products were used as templates.When 250~750 bp probe was used in CGH,1 of the 5 blastomeres was failed and another one had different karyotype from that analyzed by SRY.(2) All samples in group B and D were successfully amplified by MDA,and the non-specific bands were significantly less in the amplification of TBX1 exon 2.All 5 blastomeres were successful in CGH with the 250~750 bp probe.Moreover,the karyotype was in agreement with that of SRY.(3) When 750 ~ 2000 bp probe was used,the CGH results were suboptimal.Conclusions In WGA of single blastomere,MDA is superior to DOP-PCR in the stability and specificity.The karyotype image detected by CGH with the 250~750 bp probe is clear and homogenous.

WanfangData CO.,Ltd All Rights Reserved
About WanfangData | Contact US
Healthcare Department, Fuxing Road NO.15, Haidian District Beijing, 100038 P.R.China
Tel:+86-010-58882616 Fax:+86-010-58882615 Email:yiyao@wanfangdata.com.cn