Hi guys, i am isolating rna from whole blood using trizol method, and some of my samples have quite good A260/280 and A260/230 ratio of btw 1.7 to 2.0. In fact, i think it is necessary for me to do a rna clean up using spin column method (Qiaamp rna blood kit), but i am not sure what is the rna recovery % since i only managed to get about 10 ug of rna after trizol extraction, i afraid it will get much lesser after clean up step. FYI, i need at least 10ug for microarray work. Kindly advise. Thanks
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Dr Teeth
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Posted 29 September 2009 - 03:31 AM
wntiong, on Sep 29 2009, 12:59 AM, said:
Hi guys, i am isolating rna from whole blood using trizol method, and some of my samples have quite good A260/280 and A260/230 ratio of btw 1.7 to 2.0. In fact, i think it is necessary for me to do a rna clean up using spin column method (Qiaamp rna blood kit), but i am not sure what is the rna recovery % since i only managed to get about 10 ug of rna after trizol extraction, i afraid it will get much lesser after clean up step. FYI, i need at least 10ug for microarray work. Kindly advise. Thanks
Microarray work is much more sensitive to contaminants in your RNA sample than qRT-PCR. You should do a spin column cleanup. If your recovery is too low, consider pooling like samples.
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wntiong
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Posted 29 September 2009 - 11:02 PM
Dr Teeth, on Sep 29 2009, 07:31 PM, said:
wntiong, on Sep 29 2009, 12:59 AM, said:
Hi guys, i am isolating rna from whole blood using trizol method, and some of my samples have quite good A260/280 and A260/230 ratio of btw 1.7 to 2.0. In fact, i think it is necessary for me to do a rna clean up using spin column method (Qiaamp rna blood kit), but i am not sure what is the rna recovery % since i only managed to get about 10 ug of rna after trizol extraction, i afraid it will get much lesser after clean up step. FYI, i need at least 10ug for microarray work. Kindly advise. Thanks
Microarray work is much more sensitive to contaminants in your RNA sample than qRT-PCR. You should do a spin column cleanup. If your recovery is too low, consider pooling like samples.
I did the spin column, the band is much clearer now, is just that the amount of rna is half of the amount i obtained after trizol. How to pool my samples during spin column cleanup?
