Please help! DNA samples for whole genome sequencing

Vinn · 08-21-2016, 11:26 AM

Hi folks,

I have extracted DNA from fungi for whole genome sequencing (Truseq PCR free with 350 bp insert and HiSeq 2500 PE) using a kit. The extraction went smooth until I realized at the end that I treated my samples with only half of the recommended amount for RNase A (2 ul of 10 mg/ml instead of 2 ul of 20 mg/ml). I tried checking the DNA samples on 1% agarose and following is the gel image with 100-1000 bp ladder.

I am not sure there are RNA bands (very faint though) at around 100-200 bp. Do you think my DNA samples are good enough to go for whole genome sequencing without additional treatments? Or shall I try to treat them with RNase A again? Unfortunately, I run out of fresh samples.

I appreciate all your comments and suggestions.

Best
Vinn

jdk787 · 08-21-2016, 11:40 AM

These look fine. Even if you have some RNA in your sample it won't make into your final library after fragmentation and ligation steps.

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08-21-2016, 11:26 AM	#1
Vinn Member Location: Sweden Join Date: Nov 2014 Posts: 18	Please help! DNA samples for whole genome sequencing Hi folks, I have extracted DNA from fungi for whole genome sequencing (Truseq PCR free with 350 bp insert and HiSeq 2500 PE) using a kit. The extraction went smooth until I realized at the end that I treated my samples with only half of the recommended amount for RNase A (2 ul of 10 mg/ml instead of 2 ul of 20 mg/ml). I tried checking the DNA samples on 1% agarose and following is the gel image with 100-1000 bp ladder. I am not sure there are RNA bands (very faint though) at around 100-200 bp. Do you think my DNA samples are good enough to go for whole genome sequencing without additional treatments? Or shall I try to treat them with RNase A again? Unfortunately, I run out of fresh samples. I appreciate all your comments and suggestions. Best Vinn

08-21-2016, 11:40 AM	#2
jdk787 josh kinman Location: Austin Join Date: Apr 2014 Posts: 71	These look fine. Even if you have some RNA in your sample it won't make into your final library after fragmentation and ligation steps.