Amine-modified random primers to label probes for DNA
microarrays
Charlie C. Xiang1, Olga A. Kozhich3, Mei Chen1, Jason M. Inman3, Quang N. Phan1, Yidong Chen2
& Michael J. Brownstein1
1
Laboratory of Genetics, National Institute of
Mental Health, Bethesda, MD 20892.
2
Cancer Genetics Branch, National Human Genome
Research Institute, National Institutes of Health, Bethesda,
MD 20892.
3
SAIC Frederick, Frederick,
MD 21702.
Correspondence should be addressed to Michael J. Brownstein mike@codon.nih.gov
DNA microarrays have been used to study the expression of thousands
of genes at the same time in a variety of cells and tissues1,
2,
3. The methods most commonly used to label probes for microarray
studies require a minimum of 20 g of total RNA or 2 g of poly(A)
RNA4,
5. This has made it difficult to study small and rare
tissue samples. RNA amplification techniques and improved labeling methods have
recently been described6,
7,
8,
9. These new procedures and
reagents allow the use of less input RNA, but they are relatively
time-consuming and expensive. Here we introduce a technique for preparing
fluorescent probes that can be used to label as little as 1 g of total RNA.
The method is based on priming cDNA synthesis with random hexamer
oligonucleotides, on the 5' ends of which are bases with free amino
groups. These amine-modified primers are incorporated into the cDNA along with
aminoallyl nucleotides, and fluorescent dyes are then chemically added to the
free amines. The method is simple to execute, and amine-reactive dyes are
considerably less expensive than dye-labeled bases or dendrimers.
g of total RNA or 2 
