Size Selection Of Adaptor Ligated NGS DNA Libraries

April 04, 2018

University of Birmingham
Biosciences
Research Assistant

Overall

Quality of Results

Ease-of-Optimization

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Company:

Beckman Coulter

Product Name:

SPRIselect

Catalog Number:

B23318

Using SPRIselect reagent, DNA libraries ligated with Illumina sequencing adaptors were selected with sufficient quality and yield, and with uniform fragment libraries for sequencing.

Experimental Design and Results Summary

Application

Clean up of DNA libraries for sequencing using Illumina sequencer

Starting Material

Adaptor ligated DNA libraries

Protocol Overview

The protocol is simple and easy to follow. Follow the specific protocols mentioned with kit's instruction manual. I have used SPRIselect for clean up of 200 bp insert (fragments) ligated. First SPRIselect beads binding with DNA libraries were at 0.4X and 2nd size select was with 0.2X bead binding. Finally, libraries were washed twice with 80% ethanol and air dried. Then eluted with 20 microliter of nuclease free water. Size distribution of size SPRI size selected DNA libraries were checked by Agilent Bioanalyzer using High sensitivity DNA chip.

Tips

Thaw SPRIselect beads to room temperature at least 30 minutes before proceeding with size selection.

Results Summary

Efficient recovery of 200 bp insert size fragments after SPRIselect based clean up was observed. Agilent Bioanalyzer analysis gave fragment size distribution of 250 bp which was ideal to proceed for sequencing.

DOI or PMID #

N/A

Additional Notes

None

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Summary

The Good

Easy to follow protocol and less expensive compared to other methods.

The Bad

None

The Bottom Line

Using SPRIselect based size selection is essential practice in preparing DNA libraries for next generation sequencing. It is also possible to automate size selection if robotic equipment is present in the laboratory.

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