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Imagine that someone tries to develop a knockout mouse for a gene, but this result in lethality for the homozygous.

Is it possible to express that mutant gene only in a specific tissue of interest to get a idea of which phenotype it produces? Does someone know an experiment which used this idea or something similar?

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  • $\begingroup$ its possible, but it usually is unclear how to do it. $\endgroup$ – shigeta Apr 30 '15 at 5:15
  • $\begingroup$ @shigeta can you expand? $\endgroup$ – aaaaa says reinstate Monica Apr 30 '15 at 7:37
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    $\begingroup$ I would like to comment on the language. You start by saying "express a mutant gene only in specific tissue". Then you mention specific mutation: KO. KO-gene is lack of gene expression. At the end you again say "express that mutant gene". I suppose you meant conditional KO. $\endgroup$ – aaaaa says reinstate Monica May 1 '15 at 5:14
  • $\begingroup$ While eliminating a gene is at least understood pretty well, tissue specific expression is difficult because the DNA sequences that encode the logic in animals as to when these genes are active are poorly understood. Its been worked out only for a small number of cases I believe. $\endgroup$ – shigeta May 1 '15 at 6:41
  • $\begingroup$ @shigeta You mean that you can gene-trap tissue markers, but we don't know how certain tissue is being form or what it consists of? $\endgroup$ – aaaaa says reinstate Monica May 2 '15 at 5:31
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It is called conditional mutation.

You flox (put lox sites around) gene of interest and express Cre recombinase driven by tissue-of-interest-specific promoter.

Illustration from here:

This image shows how the Cre-lox system can be used to produce tissue-specific, GFP-marked knockout mice

Using chemically-activatable variant of Cre recombinase (cre-ER) you can create knock-out in some cells of tissue of interest, not every.

Addition: a bit weird but still useful method is to produce KO by means of CRISPR/Cas9 system. Tissue-specific expression of Cas9 protein together with gene-specific sgRNA will likely produce KO in mosaic fashion. One of positive sides is that such approach requires only one transgenic line, e.g. Tg(promoter:Cas9;sgRNA)

Addition: (via WYSIWYG's suggestion) you can express shRNA or other gene transcription-interfering product from promoter that only active in specific tissue/specific period of time. And beauty of genetic/bioengineering is that you can combine several methods for additional control, robustness, efficiency etc.

Taken from here:

Inducible and sell-specific gene KO with shRNA

Inducible and sell-specific gene KO with shRNA

For further reading:

http://cre.jax.org/introduction.html

http://link.springer.com/protocol/10.1385/1-59259-065-9:477#page-1

Update

I re-read OP question and, as commented to it, feel some confusion. So I would like to mention as well way to express mutated gene in specific tissue that, when expressed constitutively (all the time, everywhere), is lethal. Imagine some very strong oncogene. Or something like cell-toxic protein, e.g. chemically-activatable nitroreductase or light-activatable Killer-red. Or it might be mutated version of your favourite synaptic protein that, when expressed in all neurons, causes severe lethal phenotype.

What you do, is create transgenic animal with transgene-GFP fusion (nitroreductase, KillerRed, etc) that has floxed stop-codons in front, and cross that animal with transgenic with tissue-specific CreER (for example). What you get is animal that, when injected with tamoxifen, start to express mutated gene/toxic protein in tissue of interest. Then you are sure that only GFP-positive cells express that gene. That is how you "express that mutant gene only in a specific tissue of interest to get a idea of which phenotype it produces".

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    $\begingroup$ Additional info: For knockdowns, shRNA can be expressed in the desired cell type $\endgroup$ – WYSIWYG Apr 30 '15 at 5:21
  • $\begingroup$ Note that to get a knockout with Cre/LoxP system requires one further cross since F1 is heterozygous for the floxed gene. You need to cross your Cre +/0; target +/flox mice from F1 with the target flox/flox of F0 to generate Cre +/0; target flox/flox in F2. $\endgroup$ – canadianer May 1 '15 at 6:31
  • $\begingroup$ yup, that is laborious $\endgroup$ – aaaaa says reinstate Monica May 1 '15 at 6:32

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