Manufacturing DNA microarrays from unpurified PCR products
AUTOR(ES)
Diehl, Frank
FONTE
Oxford University Press
RESUMO
For the production of DNA microarrays from PCR products, purification of the the DNA fragments prior to spotting is a major expense in cost and time. Also, a considerable amount of material is lost during this process and contamination might occur. Here, a protocol is presented that permits the manufacture of microarrays from unpurified PCR products on aminated surfaces such as glass slides coated with the widely used poly(l-lysine) or aminosilane. The presence of primer molecules in the PCR sample does not increase the non-specific signal upon hybridisation. Overall, signal intensity on arrays made of unpurified PCR products is 94% of the intensity obtained with the respective purified molecules. This slight loss in signal, however, is offset by a reduced variation in the amount of DNA present at the individual spot positions across an array, apart from the considerable savings in time and cost. In addition, a larger number of arrays can be made from one batch of amplification products.
ACESSO AO ARTIGO
http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=134252Documentos Relacionados
- Rapid sequencing of unpurified PCR products by thermal asymmetric PCR cycle sequencing using unlabeled sequencing primers.
- Fluorescent dye-primer cycle sequencing using unpurified PCR products as templates; development of a protocol amenable to high-throughput DNA sequencing.
- Manufacturing DNA microarrays of high spot homogeneity and reduced background signal
- Comparing gene discovery from Affymetrix GeneChip microarrays and Clontech PCR-select cDNA subtraction: a case study
- Genome-scale design of PCR primers and long oligomers for DNA microarrays