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I am reading fw2.2: A Quantitative Trait Locus Key to the Evolution of Tomato Fruit Size (doi: 10.1126/science.289.5476.85).

The authors are trying to find the gene that makes tomatoes bigger. As far as I understand, previous research had narrowed it down to "something" in the region of the genome called fw2.2. They are trying to determine the actual genetic elements responsible (which turns out to be a gene similar to human HRAS).

Supposedly, the big-fruit allele is recessive and the small-fruit allele is dominant.

They say (in section Genetic complementation with fw2.2) that:

  1. They got a YAC containing the fw2.2.
  2. They "screened a cDNA library" with the YAC to find transcripts that were potentially relevant. (How?)
  3. They "screened a cosmid library" of genomic fragments to find the stretch of genome that the cosmid came from. (How?)

It is unclear to me how steps 2 and 3 work. Can someone explain how you can take a YAC, cDNA library and cosmid library to find the gene inside the region responsible for your trait, as these researchers describe?

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The YAC contains a segment of chromosomal DNA. By using it as a probe they could screen the cDNA library to find cDNAs derived from that genomic region. This would be done by plaque or colony hybridisation. The cDNA clones thus identified represent the "candidate genes" .

The last step is a little unclear. Clearly they are identifying cosmid clones that map to the segment of interest, but they could be using either the YAC or relevant cDNA clones (could be pooled for this step depending upon their aim) as probes. My guess is that they used individual cDNA clones to find corresponding cosmid fragments so that they could home in on the regions encoding the identified cDNAs.

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  • $\begingroup$ I am skeptical because firstly the YAC contains one allele, while the cDNA comes from an organism that has the other allele (a detail in the paper). Secondly, this becomes dubious if the gene on the ORF is spliced (like this one). For hybridizing cDNA to cosmids, there is again the potential issue of exons/introns. Or are you saying the cDNA would still hybridize if only a small part of it was complementary? $\endgroup$ – Superbest Apr 2 '14 at 7:24
  • $\begingroup$ @Superbest - hybridisation of probe to clone will occur if there is shared sequence. The fact that there might be small differences in sequence (alleles) or missing sequence (no introns in cDNA clone) will be irrelevant. Before transcriptomics this was the standard way to relate genomic DNA mapping to information about transcripts coming from cDNA cloning. $\endgroup$ – Alan Boyd Apr 2 '14 at 9:15

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