Can you please give me some suggestions and basic guidelines on the topic.

 

Thanks

can you be more specific and understandable?

It depends on many factors, but there are various strategies for making lines with lethal mutations. If you are developing a mutant for use in experiments, you can make the knockout tissue specific, for example by using recombinase sites where the recombinase is expressed in a tissue specific manner. For others, you can use a temperature-sensitive mutant, where the protein is active at low temperature but becomes inactivated at a higher temperature. You can also introduce a gene encoding shRNA with an inducible/tissue-specific promoter to knockdown expression of your target gene by RNAi.

Thanks a lot...I am mainly interested in the temperature-sensitive mutants. Can you explain the method in more details. Thanks again!

Well, you mutate a bunch of organisms (w/EMS, UV, etc.), and grow them up at room temp. Usually, temp-sens mutants are single-celled organisms, so you replate them and grow them at 37C. Whatever dies at that temperature harbors a temp-sens mutation.

Ok, first answer:

A temperature-sensitive mutant is a mutant bearing a change in amino acid (in a specific protein). This change result in the instability of a protein at high temperature.

For example, if you remove an amino acid that was allowing a solid intra-protein bound, at high temperature, the protein will be denatured because of this missing link.

 

Second, you can also create "conditional mutants". They are mutant where you need to add a reagent (an antibiotic for example) so that a gene is expressed.

 

Hope it helps.

It depends on many factors, but there are various strategies for making lines with lethal mutations. If you are developing a mutant for use in experiments, you can make the knockout tissue specific, for example by using recombinase sites where the recombinase is expressed in a tissue specific manner. For others, you can use a temperature-sensitive mutant, where the protein is active at low temperature but becomes inactivated at a higher temperature. You can also introduce a gene encoding shRNA with an inducible/tissue-specific promoter to knockdown expression of your target gene by RNAi.

 

More important to understand the contradiction between weak hydrogen bonds base pairs in DNA and codon-anticodon at termophylic bacterium. Some of them are survive at temp. above 100 C. What's about H-bonds?