Competitive genomic polymerase chain reaction
Competitive genomic polymerase chain reaction (CGP)
Variation in DNA copy number occurs in many diseases such as Down syndrome as well as in cancer. It is important to detect abnormalities in DNA copy number as associations between DNA aberrations and disease phenotype can be made and critical genes can be located.
Comparative genomic hybridization (CGH) is the standard used for genome wide analysis of DNA copy number. It is based on a two-colour fluorescence in situ hybridization (FISH). Resolution of closely spaced aberrations is not good this making it difficult to assign genomic locus. Results must also be adjusted for biases in the correlation of the heteroscedastic data distribution. Microarray based methods have higher resolution compared to CGH. However, the available arrays have gaps between probes and some regions are not available for analysis.
Competitive genomic PCR is a very useful technique to overcome the limitations. It is performed using restricted genomic DNA ligated to specific adaptors as a template. Different adaptors are added to the test and control samples and the test: reference ratio is determined by quantifying the amplified products fractionated by gel electrophoresis. In the paper, MYCN gene copy alterations in neuroblastoma- derived cells lines are being studied using CGP assay. Gene amplification in each cell line was detected. Detailed high resolution analysis by the CGP assay around the locus revealed new junctions for amplification that were not detected when a commercial array was used. It was also shown that CGP is very sensitive in detecting change in low level DNA copy number. In addition, PCR is an affordable and moderate throughput technique. It can be used to complement the hybridization technique currently used commercially for genomic analysis.
Reference:
Iwao-Koizumi K, Maekawa K, Nakamura Y, Saito S, Kawamoto S, Nakagawara A, Kato. A novel technique for measuring variations in DNA copy- number: competitive genomic polymerase chain reaction. BMC Genomics. 2007; 8: doi: 10.1186/1471- 2164- 8- 206
Loh Mun Jo-anne
(s4184551)
Variation in DNA copy number occurs in many diseases such as Down syndrome as well as in cancer. It is important to detect abnormalities in DNA copy number as associations between DNA aberrations and disease phenotype can be made and critical genes can be located.
Comparative genomic hybridization (CGH) is the standard used for genome wide analysis of DNA copy number. It is based on a two-colour fluorescence in situ hybridization (FISH). Resolution of closely spaced aberrations is not good this making it difficult to assign genomic locus. Results must also be adjusted for biases in the correlation of the heteroscedastic data distribution. Microarray based methods have higher resolution compared to CGH. However, the available arrays have gaps between probes and some regions are not available for analysis.
Competitive genomic PCR is a very useful technique to overcome the limitations. It is performed using restricted genomic DNA ligated to specific adaptors as a template. Different adaptors are added to the test and control samples and the test: reference ratio is determined by quantifying the amplified products fractionated by gel electrophoresis. In the paper, MYCN gene copy alterations in neuroblastoma- derived cells lines are being studied using CGP assay. Gene amplification in each cell line was detected. Detailed high resolution analysis by the CGP assay around the locus revealed new junctions for amplification that were not detected when a commercial array was used. It was also shown that CGP is very sensitive in detecting change in low level DNA copy number. In addition, PCR is an affordable and moderate throughput technique. It can be used to complement the hybridization technique currently used commercially for genomic analysis.
Reference:
Iwao-Koizumi K, Maekawa K, Nakamura Y, Saito S, Kawamoto S, Nakagawara A, Kato. A novel technique for measuring variations in DNA copy- number: competitive genomic polymerase chain reaction. BMC Genomics. 2007; 8: doi: 10.1186/1471- 2164- 8- 206
Loh Mun Jo-anne
(s4184551)
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