I get that you're cloning a whole cDNA library, but why does E. coli need a complementary gene to allow the gene expression? Why is that complementary gene considered to be a mutant? AND how does one detect the gene of interest after cloning?

What is this kind of cloning used for?


closed as unclear what you're asking by AliceD, another 'Homo sapien', Bryan Krause, David, James Feb 23 '17 at 3:14

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  • $\begingroup$ Your question is unclear, which is why there are four votes to close it. I have constructed cloned cDNA libraries and I have no idea what your question means. Please clarify it if you want an answer. $\endgroup$ – David Feb 22 '17 at 19:41
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    $\begingroup$ I thought your question was clear enough, but it may be closed as too broad since there are multiple questions on what is a rather large subject. Given that you apparently now understand the basics, perhaps you could narrow your question to the specifics of what is still confusing you. Or start a new question. $\endgroup$ – canadianer Feb 22 '17 at 20:36
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    $\begingroup$ PS You're not "too undergrad" (ha!) for the site. Most questions asked here are at an undergrad or popular science level. $\endgroup$ – canadianer Feb 22 '17 at 20:38
  • $\begingroup$ @canadianer great that makes me feel much better because I really love this site! I will take you up on the advice to narrow the questions, thanks! $\endgroup$ – Kirstie Alexandra Jiles Feb 22 '17 at 20:59

I figured it out. You're complementation cloning when you know the function of a mutant sequence but not it's sequence. For example, your organism (human) gets skin cancer from too much UV light exposure.

With complementation, two mutants can restore the wild type phenotype (well functioning UV repair).

We know E. coli has a well studied UV repair gene so you transform the cDNA library from your human into an E. coli which we know has the mutation, then grow them all under intense UV light. The E. coli with the human mutant allele (the right cDNA fragment) will survive, so you can amplify that colony, sequence it, and then use it as a diagnostic tool in humans.

I'm wondering why that was vague. Is "complementation cloning" not a well used name for that technique? Was the question too broad or elementary? It's the first time I've used Stack and it seems I may be a little too undergrad for this crowd.

  • $\begingroup$ If my answer clarifies, please feel free to fill in gaps I left out because I'm still trying to understand some of the reasons this technique works and why we'd use it. $\endgroup$ – Kirstie Alexandra Jiles Feb 22 '17 at 20:07

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