A method for generating selective DNA probes for the analysis of C-negative regions in human chromosomes.
| Autor: | Morozkin ES; Institute of Chemical Biology and Fundamental Medicine, Siberian Division, Russian Academy of Sciences, Novosibirsk. morozkin @ niboch.nsc.ru, Loseva EM, Karamysheva TV, Babenko VN, Laktionov PP, Vlassov VV, Rubtsov NB |
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| Jazyk: | angličtina |
| Zdroj: | Cytogenetic and genome research [Cytogenet Genome Res] 2011; Vol. 135 (1), pp. 1-11. Date of Electronic Publication: 2011 Jul 28. |
| DOI: | 10.1159/000330124 |
| Abstrakt: | Linker-adapter polymerase chain reaction (LA-PCR) is among the most efficient techniques for whole genome DNA amplification. The key stage in LA-PCR is the hydrolysis of a DNA sample with restriction endonucleases, and the choice of a restriction endonuclease (or several endonucleases) determines the composition of DNA probes generated in LA-PCR. Computer analysis of the localization of the restriction sites in human genome has allowed us to propose an efficient technique for generating DNA probes by LA-PCR using the restriction endonucleases HaeIII and RsaI. In silico hydrolysis of human genomic DNA with endonucleases HaeIII and RsaI demonstrate that 100- to 1,000-bp DNA fragments are more abundant in the gene-rich regions. Applying in situ hybridization to metaphase chromosomes, we demonstrated that the produced DNA probes predominantly hybridized to the C-negative chromosomal regions, whereas the FISH signal was almost absent in the C-positive regions. The described protocol for generating DNA probes may be successfully used in subsequent cytogenetic analysis of the C-negative chromosomal regions. (Copyright © 2011 S. Karger AG, Basel.) |
| Databáze: | MEDLINE |
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