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Describe a technique for using next-generation sequencing to estimate how many mature mRNA transcripts have been...

Describe a technique for using next-generation sequencing to estimate how many mature mRNA transcripts have been made (including how you have decided to make sure you will only sequence mature mRNA, and not other RNAs)?

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Answer #1

Technique involves :

isolation of total RNA from the cell.

For isolation of mRNA only, we can use its special feature that is poly A tail. We can purify all mRNA by affinity chromatography using poly T oligonucleotides as the stationary phase.

After isolation of mRNA from the cell, make cDNA of mRNAs using poly A abs primer.

Add linkers to the flanking regions of these cDNA regarding the primers to be used in sequncing.

Then sequencing is done. After sequence analysis, alignments, we can determine the number of mRNA made in the cell.

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