TruSeq small RNA final products Peaks

shahideh · 03-08-2013, 10:26 AM

Hi everyone,

I have tried to make small RNA libraries from insect RNAs and I should say that it was my first experience. I used TruSeq small RNA kit. I have attached my final products (samples 1-3, 4 is just water) run in Experion DNA 1K chip from Bio-Rad after gel purification and ethanol precipitation. I appreciate if you guys let me know if you think they look good to send for sequencing.
My ultimate goal is discovering known and novel viruses in insect. So, I am looking for any kind of small RNAs (18-30 nt).

Thanks

kcchan · 03-10-2013, 10:03 PM

Your libraries look perfect! The goal of the gel cut is to remove the primer and adapter dimer bands at ~125 and 135bp while keeping your small RNA libraries, which in your case are ~153-165bp. Looking from these traces you have nothing but the library peak. The yields also look good enough for pooling and sequencing.

shahideh · 03-12-2013, 09:19 AM

Thanks so much for your reply

F2000 · 03-14-2013, 09:02 AM

Hi,
Which method did you use for isolating the small RNA fragment from the adapter dimer.

Thanks

shahideh · 03-14-2013, 10:06 AM

HI F2000,

I did gel purification. I followed the TruSeq protocol

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03-10-2013, 10:03 PM	#2
kcchan Senior Member Location: USA Join Date: Jul 2012 Posts: 184	Your libraries look perfect! The goal of the gel cut is to remove the primer and adapter dimer bands at ~125 and 135bp while keeping your small RNA libraries, which in your case are ~153-165bp. Looking from these traces you have nothing but the library peak. The yields also look good enough for pooling and sequencing. Last edited by kcchan; 03-10-2013 at 10:05 PM. Reason: Derp, didn't read the initial post fully. Sample 4 is NTC.

03-12-2013, 09:19 AM	#3
shahideh Junior Member Location: US Join Date: Feb 2013 Posts: 8	Thanks so much for your reply

03-14-2013, 09:02 AM	#4
F2000 Junior Member Location: France Join Date: Mar 2013 Posts: 1	Hi, Which method did you use for isolating the small RNA fragment from the adapter dimer. Thanks

03-14-2013, 10:06 AM	#5
shahideh Junior Member Location: US Join Date: Feb 2013 Posts: 8	HI F2000, I did gel purification. I followed the TruSeq protocol