Question: Difference between Rad-Seq and amplicon seq
0
gravatar for CY
12 weeks ago by
CY40
United States
CY40 wrote:

As far as I know, both Rad-Seq and amplicon sequencing are aim to interrogate specific regions of the genome by cutting the region with restriction enzymes or primers.

Can anyone share some difference between these two techniques in any aspect, like goal, technical limitation or......? Really appreciate!

ADD COMMENTlink written 12 weeks ago by CY40

RADseq does not aim to target specific regions of the genome, the intention is to sample the genome randomly, to sequence just a fraction of it and cut down costs.

ADD REPLYlink written 12 weeks ago by h.mon9.0k

Then what is the purpose of this technique?

If it is just random cutting, the purpose seems to be looking for any SNP or QTL within any regions. But you are not covering all the regions using this technique and you will probably missing lots of candidates. It seems not reasonable to me...

ADD REPLYlink written 12 weeks ago by CY40

Then what is the purpose of this technique?

RADSeq: next-generation population genetics

ADD REPLYlink written 12 weeks ago by h.mon9.0k

I read the paper and still felt little difference between capture based sequencing and Rad-Seq. Both are capturing region near which restriction enzyme which specific sequencing bind to.

ADD REPLYlink written 12 weeks ago by CY40

I read that RAD-seq aimes to reduce the genome complexity by using restriction enzymes which cut the known regions in genomes so instead of sequencing whole of a complex genome we just sequence the genome in the border of restriction sites.

ADD REPLYlink written 12 weeks ago by Fereshteh2.7k

I can image it is cost-effective to selecting only the region near which the restriction enzymes bind and get high depth for population study. The questions is: Is the region selection (restriction enzyme selection) is entirely random? Or there are some sequence selection through the design of restriction enzyme?

ADD REPLYlink written 12 weeks ago by CY40
1

A standard RADseq project sequences all regions adjacent to, say, the SbfI cut site. That may be 10,000 or more loci across a genome, but also could be just 0.1% of the genome. It is very difficult to choose a restriction enzyme or combination of enzymes that would sequence more than one SNP of interest, so it is not useful for targeted sequencing. If you just have a SNP or two, other methods are more specific and cheaper. If you have many particular SNPs, it cannot be done.

What it does well is sequence those 10,000 loci consistently in many samples, allowing a population to be studied for the cost of a single lane of a sequencer. Having several thousand SNPs in a non-model organism allows many population analyses to be done.

ADD REPLYlink written 12 weeks ago by SNPsaurus40

Hi SNPsaurus

Thanks for explaining.

However, what do you mean "It is very difficult to choose a restriction enzyme or combination of enzymes that would sequence more than one SNP of interest" and "Having several thousand SNPs in a non-model organism allows many population analyses to be done"? It is kind of conflicting sentence to me.

Also, I image the selection of restriction enzyme (covered genome region) is not entirely random. Is there any criteria used in this?

ADD REPLYlink written 12 weeks ago by CY40
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