Genet Matthew D, Cartwright Ian M, Kato Takamitsu A
Department of Environmental and Radiological Health Sciences, Colorado State University, 1618 Campus Delivery, Fort Collins, CO 80523, USA.
Mol Cytogenet. 2013 Oct 8;6(1):42. doi: 10.1186/1755-8166-6-42.
Fluorescence in situ Hybridization (FISH) utilizes peptide nucleic acid (PNA) probes to identify specific DNA sequences. Traditional techniques have required the heat denaturing of the DNA in formamide followed by multiple hours at moderated temperatures to allow the probe to hybridize to its specific target. Over the past 30 years, advancements in both protocols and probes have made FISH a more reliable technique for both biological research and medical diagnostics, additionally the protocol has been shortened to several minutes. These PNA probes were designed to target and hybridize to both DNA and RNA, and PNA-protein interactions still remain unclear.
In this study we have shown that a telomeric single stranded specific PNA probe is able to bind to its target without heat denaturing of the DNA and without formamide. We have also identified a centromere specific probe, which was found to bind its target with only incubation with formamide.
Certain PNA probes are able to hybridize with their targets with minimal to no denaturing of the DNA itself. This limited denaturing preserves the chromosome structure and may lead to more effective and specific staining.
荧光原位杂交（FISH）利用肽核酸（PNA）探针来识别特定的DNA序列。传统技术需要在甲酰胺中对DNA进行热变性处理，然后在适度温度下孵育数小时，以使探针与其特定靶标杂交。在过去30年中，方案和探针方面的进展使FISH成为生物学研究和医学诊断中更可靠的技术，此外，该方案已缩短至几分钟。这些PNA探针被设计用于靶向并与DNA和RNA杂交，而PNA与蛋白质的相互作用仍不清楚。
在本研究中，我们表明端粒单链特异性PNA探针能够在不使DNA热变性且不使用甲酰胺的情况下与其靶标结合。我们还鉴定出一种着丝粒特异性探针，发现其仅在与甲酰胺孵育时就能与其靶标结合。
某些PNA探针能够在DNA本身极少变性或不变性的情况下与其靶标杂交。这种有限的变性保留了染色体结构，并可能导致更有效和特异的染色。
