Optimized amplification and fluorescent labeling of small cell samples for genomic array-CGH.
BACKGROUND
Whole genome amplification (WGA) is usually needed in the genetic analysis of samples containing a low number of cells. In genome-wide analysis of DNA copy numbers by array comparative genomic hybridization (array-CGH) it is very important that the genome is evenly represented throughout the amplified product. All currently available WGA techniques are generating some degree of bias.


METHODS
A way to compensate for this is using a reference sample which is similarly amplified, as the introduced amplification bias will be leveled out. Additionally, direct labeling of the amplified DNA is performed to bypass the currently widely applied random primed labeling, which involves an additional amplification of the product and is introducing extra bias.


RESULTS
In this article it is shown that equal processing of the test and reference sample is indeed crucial to generate an optimal array-CGH profile of amplified DNA samples. Also presented here is that the labeling method may significantly effect the array-CGH result, it is shown that with direct chemical labeling using platinum derivates (ULS labeling) optimal array-CGH results are obtained.


CONCLUSIONS
We show that an optimized WGA strategy for both test and reference sample in combination with direct chemical labeling results in a reliable array-CGH profile of samples as low as a 30 cell equivalent.
Knijnenburg
Jeroen
J
Department of Molecular Cell Biology, Leiden University Medical Center, Leiden, The Netherlands.
van der Burg
Marja
M
Tanke
Hans J
HJ
Szuhai
Károly
K
D045744:Cell Line, Tumor; D005456:Fluorescent Dyes / Q000032:analysis* / Q000378:metabolism; D015894:Genome, Human / Q000235:genetics*; D006801:Humans; D008297:Male; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D009693:Nucleic Acid Hybridization / Q000379:methods*; D016133:Polymerase Chain Reaction; D013194:Staining and Labeling
D005456:Fluorescent Dyes