Genetic variation in the lab (V0358)
David Ackerley of Victoria University, Wellington, explains the techniques used in his lab to make variations of a particular gene for directed evolution.
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Scientists use a technique called PCR to make exact copies of a gene. In David Ackerley’s lab, they use different types of PCR to make copies of the gene that have random variations in the DNA sequence. The researchers then test to see whether these random variations are good or bad for the function of the protein the gene codes for.
David Ackerley (Victoria University, Wellington): We have a number of tricks to introduce high levels of mutation into just a gene, and I think students at year 12 and year 13 are familiar with a process called PCR, which is a way of artificially replicating DNA in a test tube. We actually use that same process to introduce errors into our gene. The more random you can make these things, then the more variation you are going to get, and you get a bigger pool of variation to apply your selective pressure to.
Se we use these ‘sloppy’ forms of PCR to introduce mutations into our genes of interest. We use other forms of PCR to try and mimic the sexual recombination process – so, snipping up and shuffling those genes and stitching them back together. But, essentially, it’s the same deal – it’s just getting variation into a set of genes that you start with.
So we have no sensible way of predicting which changes might improve the activity that we're after. And that's why taking this random approach of just sprinkling random mutations in there and letting nature take its course, and selecting for the ones that have the improvement that we want, allows us to achieve that outcome without knowing exactly which changes need to be made in the first place.
- 05 December 2008
- The University of Waikato