Bacterial Genome Sequencing Using MiSeq Reagent Kit V3

University of Birmingham
School of Biosciences
Postdoctoral Research Fellow

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Quality of Results

Ease-of-Optimization

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Company:

Illumina

Product Name:

MiSeq Reagent kit v3

Catalog Number:

MS-102-3001

I have used Illumina's MiSeq Reagent Kit V3, which is a reagent cartridge containing pre-filled reagents for cluster generation, sequencing primers during sequencing chemistry which takes place in MiSeq sequencing platform E. coli isolates genome sequencing. 16 pM denatured library was loaded into the sample well in the reagent cartridge and then the cartridge was loaded onto MiSeq. I got 18 million reads with a Q score of greater than 30.

Experimental Design and Results Summary

Application

Bacterial genome sequencing in MiSeq platform

Starting Material

16 pM bacterial DNA libraries prepared using Nextera XT DNA library prep kit

Protocol Overview

Thaw reagent cartridge to room temperature using room temperature water in a trough. Then mix well contents of the reagent cartridge by shaking horizontally. Next load the denatured 16 pM equally pooled DNA library sample into designated sample loading well. Then place the cartridge in the Miseq and follow on-screen instructions in MiSeq to start sequencing reaction.

Tips

None

Results Summary

Got 18 million reads with Q scores for bases greater than 30. Run specifications were paired end 150 bp sequencing.

Additional Notes

None

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Summary

The Good

Easy-to-follow protocol. Takes less time to perform the experiment.

The Bad

None

The Bottom Line

We routinely use this reagent kit for bacterial genome sequencing and targeted amplicon sequencing as well as for transposon sequencing.

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