I'm facing problems regarding extraction of RNA from corneal tissues acquired post DSEK. I have been using Qiagen RNA extraction Kit so far. But the conc. is as low as 2ng/ul with 260/280 2.23 and 260/230 1.86. Does anybody have any better method(s) or kit to extract RNA from such tissues?
With corneas you will always get pretty low amts of RNA compared to other tissue. I used to work with mouse corneas with tiny amts of RNA. Are you using an RNAEasy or Qiazol based extraction? Qiazol is essentially equivalent to Trizol and can often give a better yield of RNA especially when dealing with tough tissue with a relatively low RNA content to start with.
TRIZOL method is the best method to isolate RNA , But you please sure about all the things to be double auto-claved and taking care when we are collecting supernatent, and use TRIZOL immediately after thawing, dont wait for so much time.
All the Best
Thank you Anilkumar, Ian and Aneesha for your expert advise. I was skeptic to use Trizol method for its harsh way of RNA extraction. But now I feel I should atleast try Trizol with the control corneas.
in my opinion the best method to isolate RNA from cornea is the RNeasy Plus Micro kit of Qiagen. The amounts of RNA from cornea are always lower than in others tissues. It is very important introduce the cornea tissue after the DSAEK surgery in RNA protect buffer to protect the RNA and avoid his degradation
Hi Gargi. Have you extracted so far a pure RNA from corneal tissue? I am facing problems in RNA extraction using Trizol and would appreciate your assistance.
Dear Waseem, I am using RNeasy micro kit. It is specially equipped for very small tissue sections. Fresh tissues are always better to expect a good yield (10-15ng/ul).
All the best.
https://www.qiagen.com/in/shop/sample-technologies/rna-sample-technologies/total-rna/rneasy-micro-kit/
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