Abstract
Comparative genomic hybridization (CGH) to metaphase chromosomes has
been widely used for the genome-wide screening of genomic imbalances
in tumor cells. Substitution of the chromosome targets by a matrix
consisting of an ordered set of defined nucleic acid target sequences
would greatly enhance the resolution and simplify the analysis procedure,
both of which are prerequisites for a broad application of CGH as
a diagnostic tool. However, hybridization of whole genomic human
DNA to immobilized single-copy DNA fragments with complexities below
the megabase pair level has been hampered by the low probability
of specific binding because of the high probe complexity. We developed
a protocol that allows CGH to chips consisting of glass slides with
immobilized target DNAs arrayed in small spots. High-copy-number
amplifications contained in tumor cells were rapidly scored by use
of target DNAs as small as a cosmid. Low-copy-number gains and losses
were identified reliably by their ratios by use of chromosome-specific
DNA libraries or genomic fragments as small as 75 kb cloned in P1
or PAC vectors as targets, thus greatly improving the resolution
achievable by chromosomal CGH. The ratios obtained for the same chromosomal
imbalance by matrix CGH and by chromosomal CGH corresponded very
well. The new matrix CGH protocol provides a basis for the development
of automated diagnostic procedures with biochips designed to meet
clinical needs.
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