Library Preparation Troubleshooting

Library preparation will likely define the quality of your sequencing data. While the construction of a genomic library will be slightly different from a cDNA library, with different kits for each, some basic principles apply to both. Despite this, things can still go awry.

If this is the case for you, have no fear! This ZAGENO troubleshooting guide is here to help.

One Marketplace. All of Biotech.
Your Perfect Solution - Millions of Products - Expert Advice

Poor Yield

  • Poor DNA quality: Make sure that your material is not degraded and has been stored correctly

  • Insufficient purity: Be sure to purify your DNA preparation before using it for library creation

  • Wrong quantification: The quantification of your DNA might not have been executed precisely enough. Check the quality of your quantification

  • Insufficient amplification: Add two or three amplification steps in your protocol

  • Insufficient adapter ligation Make sure that your ligation followed the supplier protocol

Uneven Coverage

  • Too may amplification cycles: Excessive amplification can amplify biased sequence coverage. Reduce the amplification steps in your protocol

  • Amplification range over instrumental sensitivity: The amplification might have pushed some sequences over the instrumental sensitivity of your instrument. Reduce the amplification steps in your protocol.

If you plan on building a new DNA library, follow these tips to help minimize mistakes

  • Use short DNA fragments

  • Purify your DNA to remove small fragments

  • Check for the presence of adapters. This can be performed by PCR or by gel electrophoresis.