Scanning copy number and gene expression on the 16p13.3-13.2 chromosomal region by the systematic multiplex polymerase chain reaction and reverse transcription-polymerase chain reaction methods.
We developed the systematic multiplex reverse transcription-PCR (SM RT-PCR) method that is distinguishable from other multiplex RT-PCR methods by optimized PCR conditions allowing amplification of sequences that fall within a single exon of genes of similar band intensity using genomic DNA template as a calibration standard. Using an SM RT-PCR system of proto-oncogenes and tumor suppressor genes, we previously showed that the SM RT-PCR system, which was developed for cDNA expression analysis, could also be used for a more exquisite analysis of copy number changes in genomic DNA. Here we report that the SM PCR method semiquantitatively detected less than a two-fold difference in copy number. Furthermore, we also report the results of subchromosomal scanning of copy number and expression using the SM PCR and SM RT-PCR methods. We identified and characterized the novel homozygous deletion that spans over 12-plus genes on 16p13.3-13.2 in the MDA-MB-468 breast cancer cell line.
Yamamoto
Miyako
M
Cancer Genetics and Epigenetics Program, Burnham Institute for Medical Research, La Jolla, CA 92037, USA. fyamamoto@burnham.org
Ahn
Ray Hyungjoo
RH
Yamamoto
Fumiichiro
F
D002885:Chromosomes, Human, Pair 16 / Q000235:genetics*; D005260:Female; D018628:Gene Dosage; D015870:Gene Expression; D006801:Humans; D008297:Male; D016133:Polymerase Chain Reaction / Q000379:methods*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D017422:Sequence Analysis, DNA