Checking Adaptor Ligated Whole Genomic DNA Sequencing Library Fragment Size

May 03, 2018

University of Birmingham
Biosciences
Research Assistant

Overall

Quality of Results

Ease-of-Optimization

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Company:

Agilent Technologies

Product Name:

High Sensitivity DNA Reagents

Catalog Number:

5067-4626

I have sheared whole genomic DNA sequencing library to 400 bp and ligated them with Illumina sequencing adaptors. After ligation and clean up, I have checked the size of the library using Agilent's High Sensitivity DNA reagents with high sensitivity DNA chip in a Agilent Bionalyzer 2100. All libraries checked were of 480 bp size, which is expected and ideal to proceed for sequencing.

Experimental Design and Results Summary

Application

Whole genome sequencing library size determination, NGS

Starting Material

5 ng of adaptor ligated and index attached whole genomic DNA libraries

Protocol Overview

The protocol very simple. Follow the easy instructions which come with High Sensitivity DNA Reagents kit for preparing reagents and loading onto high sensitivity DNA chip. After loading reagents and samples onto the high sensitivity DNA chip, insert it into Agilent Bioanalyzer and perform run.

Tips

Avoid generating air bubbles while loading gel-dye matrix and also while loading DNA samples into the wells of high sensitivity DNA chip

Results Summary

I have test 8 sequencing DNA libraries. All the libraries were of 480 bp in seize and was ideal for my libraries to proceed for sequencing.

DOI or PMID #

N/A

Additional Notes

None

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Summary

The Good

Easy to follow protocol and performed within 45 minutes.

The Bad

Bit expensive. You can't just run 2 or 3 DNA libraries with DNA chip having slots for 11 samples, where in other 8 wells goes unused and the same chip can't be used again.

The Bottom Line

This is a must do quality check and hence it (High sensitivity DNA reagents kit) is required in NGS lab. I suggest using this reagent kit for checking DNA library size.

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