Hi,
My customer increased the primer concentration and got an additional peak (marked with the red square below) near the main peak. Would you kindly let me know what is the possible reason? Please see the image below. Thank you so much!
Hi,
My customer increased the primer concentration and got an additional peak (marked with the red square below) near the main peak. Would you kindly let me know what is the possible reason? Please see the image below. Thank you so much!
Hello,
While we do have a general idea of what might be causing this additional peak, we would be better able to support your customer if you could send the original data file (.mxp) in which these results were observed to qpcr@agilent.com<mailto:qpcr@agilent.com>.
Could you please forward this file?
Thank you,
Albert Grafsky
Manager,
Technical Services
Agilent Technologies, Inc.
T: +1 800 227 9770 | M: +1 858 229 1118 | www.agilent.com<http://www.agilent.com/>
F: +1 858 597 0619 | al.grafsky@agilent.com
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Before seeing the original .mxp file, one comment that can be made is that higher temperature dissociation peaks such as the one seen here have frequently been associated with the presence of rare splice variants of the target sequence. Alternatively non-specific product formation, due to the primers hybridizing to sequence other than the intended target, has also been known to cause such peaks. Since you indicated that this peak appeared after the customer increased their primer concentration, it seems that the second cause is the more likely.
Before seeing the original .mxp file, one comment that can be made is that higher temperature dissociation peaks such as the one seen here have frequently been associated with the presence of rare splice variants of the target sequence. Alternatively non-specific product formation, due to the primers hybridizing to sequence other than the intended target, has also been known to cause such peaks. Since you indicated that this peak appeared after the customer increased their primer concentration, it seems that the second cause is the more likely.