In molecular genetics, I read, some hybridization assays, the hybridization stringency is deliberately designed to be low; other hybridization assays depend on very stringent hybridization. What is meant by hybridization stringency, how can high and low stringency be achieved, and in what circumstances would very high and low hybridization stringencies be required for a hybridization assay?
Best wishes
Hi,
the main factor for hybridization stringency is a comlementarity of bases followed by temperature and time of hybridization hand in hand with type of buffer (pH).
hybridisation depends on the bases on the probe and the hydrogen bonding between the bases on the target and on the probe matching. A high stringency hybridisation is when the hybridisation conditions....sequence homology,salt concentration and temperature all combine to allow only the perfect homology probe to anneal to its target . This is needed when gene specific probes are used and there may be similar sequences in other genes that must not be allowed to confuse the result. High stringency is caused by high temperature, low salt and perfect homology of the probe. Low stringency is high salt and low temperature which stabilise base binding and possibly low homology of probe .
High stringency conditions also gove a cleaner background signal.
Low stringency is used to prove binding has occurred before doing a high stringency wash or if the target sequence is uncertain ( polymorphism or gene families) and you want to make sure of binding accepting a loss off stringency and possibly accuracy
Dear Seyed
The nucleic acid hybridization used the labeled probe to label matching sequences by base-pairing, and the unbound probe is removed by a series of stringency washes.
At high stringency, duplexes form only between strands with perfect one-to-one complementarity, while lower stringency allows annealing between strands with some degree of mismatch between bases.
Good Luck
Dear Seyed,
Check this link you may get what's you want.
https://www.ndsu.edu/pubweb/~mcclean/plsc731/dna/dna6.htm
Regards
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