Using the TruSeq® Nano DNA, I missed my first Illumina bank (no DNA with bioanalyser).
Can you share your experience, it can help me, because I don't know which step I missed.....
Out of 8/9 steps you might have followed before going for the Bioanalyser quantification. Are these follows
Sample Prep Workflow
Prepare Adapter Setup
Fragment DNA
Perform End Repair and Size Selection
Adenylate 3' Ends
Ligate Adapters
Enrich DNA Fragments
Validate Library
Normalize and Pool Libraries
As per my experience, Bioanalyser itself will give wrong result when the ladder lane itself not run perfectly, you will get result as nill.
Most probably we might have miss to do Enrichment of DNA fragments or while doing library purification step using Ampure kit, we may miss it due to in appropriate duration of incubation as well as protocol not followed properly.
These are the some of errors could be happen manually.
Regards
Murugan N
Dear Murugan,
I try to make my DNA library twice, without any DNA at the end....
Then I try it using another kit, and it work!s
Definitely, I think this is a problem with my ligase enzym. I think that it did not work with my first kit, I don't know why!
As the cloning, it seems that this step is very sensitive.
Ya, There may be problem with the kits also possible.,
Some times if the reagents kept above it optimal temperature, NGS reagents are very sensitive to Temperature, They must be always at its optimal temperature (-20 0c or 4 o c.
So keep your reagents at optimal storage temperature, just take it away 5 mins before going to use it and replace it at a time of 5 mins.
If the reagent using at first time itself , if not working properly na, u could ask manufacturer to replace.
I wish u all success.
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