Hi,
Referring to the above matter, I would like to know the contribution of the CAG repeats length to the risk of developing prostate cancer. In this regards, I am interested in counting accurately the CAG repeats length in a DNA sequence. I have extracted DNA from the buffy coat obtained from prostate cancer patients' blood. The DNA was then amplified using PCR and subjected to gel electrophoresis using a specific primer set. The band was cut before sending to DNA sequencing.
The DNA sequence generated with the forward primer is as attached. Based on the DNA sequence, I used ((CAG)nCAA) method as suggested by Hsing et al (2000) "Polymorphic CAG and GGN Repeat Lengths in the Androgen Receptor Gene and Prostate Cancer Risk: A Population-based Case-Control Study in China" to count the CAG repeat length. I wonder whether CAG repeat more than one, for example, CAGCAG which lies outside of the main CAG repeats length is also considered as CAG repeat? And, are there any other methods to count the CAG repeats length?
Hope to hear from you all soon.
Thank you so much.
Best regards,
Seng
Hi there,
CAG repeats belongs to a group of short simple sequence stretches which is thought to arise by slippage like events randomly occurring internal repetitive sequence stretches, they are generally hypervariable in length, which are interspersed with a unique triplets, like CAG withCAA - (CAG)n CAA. Therefore, a piece of CAGCAG sequence , if it is flanked with other nucleotide composition rather than an unique triplets, would not be considered as CAG repeat, it is just a fragment of genomic DNA .
Decades ago, people had used SSLP ( simple sequence length polymorphisms) to identifying short repetitive elements and genome mapping, and now RFLA ( Restriction Fragment Length Polymorphisms) probes are frequently used in genome mapping and variation analysis .
Hope it would help !
Hi there
Just like what other colleagues mentioned above, its not the best way to use sequencing for the determination of the number of CAG repeats , however its okay to use it for confirming the fragment analysis results if needed. CAG repeats outside of the main repetitive region should not be considered.
Good luck
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