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本帖最后由 细胞海洋 于 2012-3-26 13:34 编辑
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8 i3 k: Z/ i5 P: h8 b. gUse of Comet-FISH in the study of DNA damage and repair:( f4 Y/ W- J# G- U4 G& j4 P
The Comet-FISH technique is a useful tool to detect overall and region-specific DNA damage and repair in individual cells. It combines two
" d/ T' P. i, P* lwell-established methods, the Comet assay (single cell gel electrophoresis) and the technique of fluorescence in situ hybridization (FISH). Whereas
2 _; t' P$ u: X4 s8 C5 r# K& o8 sthe Comet assay allows separating fragmented from non-fragmented DNA, FISH helps to detect specifically labelled DNA sequences of interest,
4 Y0 b2 K6 I- Qincluding whole chromosomes. Thus the combination of both techniques has been applied in particular for detection of site-specific breaks in DNA
+ d) P# C) K6 l8 H% E& Jregions which are relevant for development of different diseases. This paper reviews the relevant literature and presents three examples on how
' f F& f4 z9 A$ Q/ OComet-FISH was used for studying the induction of DNA damage by genotoxic compounds related to oxidative stress in colon cancer-relevant
' Q) k1 W' A6 {) A1 t) m1 ugenes (TP53, APC, KRAS) of a colon adenoma cell line. The accumulated evidence on relative sensitivity of these genes in comparison to global
: i1 P/ c J& ?/ ]damage allows a more definite conclusion on the possible contribution of the genotoxic factors during colorectal carcinogenesis. Telomere fragility/ L8 t/ [$ C5 f2 |' ^- s9 i3 \/ h+ m
was compared in different cell lines treated with cytostatic agents, and revealed new patterns of biological activities through the drugs and different) Y+ s" p9 o9 Z/ @$ J
sensitivities of the cell lines that were found to be associated with their tumour origin. A third example relates to measuring repair of specific gene* `; i3 i# Z c- _; @: q$ M
regions using Comet-FISH, a method that can be developed to biomarker application. Taken together, available data suggests that Comet-FISH& N3 @3 J% i7 W% m+ S
helps to get further insights into sensitivity of specific DNA regions and consequently in mechanisms of carcinogenesis. Although the nature of the
4 O* m- q5 O. f O: T- [1 Tmeasured Comet-FISH endpoint precludes us from stating basically that damage and repair are occurring within the specific gene, it is at least: K2 T8 f; @' [7 ?: I: O& G* q
possible to evaluate whether the damage and repair are occurring within the vicinity of the gene of interest.
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发表于 2012-3-26 10:54
