about cDNA library

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xyfwht
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about cDNA library

Post by xyfwht » Tue Jan 19, 2010 5:21 am

when we construct cDNA library about leaves inoculated by pathogen,we should isolate and purify the mRNA,why can't we directly use the total RNA ? is it has a great effects on result? :? thank you!

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biohazard
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Post by biohazard » Tue Jan 19, 2010 7:45 am

Well, complementary DNA is by default product of messenger RNA, so if you got lots of other RNA molecules hanging around you may also synthesize bits and pieces of non-coding DNA and then its not quite cDNA library. Of course, to some degree this may happen when working with mRNA as well. The biggest effect is the overall poorer quality of your cDNA library and bigger use of reagents when making it. But I think you can also make a shomehow functional cDNA library also out of total RNA; after all, most of the other RNA around there shouldn't even give you DNA products because their conformation does not allow reverse transcriptase to work.

It is just that there are many nifty kits for isolating mRNA, so why would you want to use total RNA? :)

xyfwht
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Re:

Post by xyfwht » Wed Jan 20, 2010 8:54 am

biohazard wrote:Well, complementary DNA is by default product of messenger RNA, so if you got lots of other RNA molecules hanging around you may also synthesize bits and pieces of non-coding DNA and then its not quite cDNA library. Of course, to some degree this may happen when working with mRNA as well. The biggest effect is the overall poorer quality of your cDNA library and bigger use of reagents when making it. But I think you can also make a shomehow functional cDNA library also out of total RNA; after all, most of the other RNA around there shouldn't even give you DNA products because their conformation does not allow reverse transcriptase to work.

It is just that there are many nifty kits for isolating mRNA, so why would you want to use total RNA? :)




thank you very much! when i constructed the cDNA library ,i neglected the isolation of mRNA.now ,should i do it again?
i'm interesting in the functional cDNA library that you had mentioned,can you elaborated further on it? thank you again! :D

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