Open AccessCapillary Electrophoresis-Based Functional Genomics Screening to Discover Novel Archaeal DNA Modifying Enzymes
Author(s) -
Kelly M. Zatopek,
Samantha L. Fossa,
Katharina Bilotti,
Paul J. Caffrey,
Léa Chuzel,
Alexandra M. Gehring,
Gregory J. S. Lohman,
Christopher H. Taron,
Andrew F. Gardner
Publication year - 2022
Publication title -
applied and environmental microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.552
H-Index - 324
eISSN - 1070-6291
pISSN - 0099-2240
DOI - 10.1128/aem.02137-21
Subject(s) - fosmid , genomic dna , capillary electrophoresis , nucleic acid , biology , computational biology , dna sequencing , genomics , massive parallel sequencing , dna , genomic library , metagenomics , functional genomics , illumina dye sequencing , genetics , microbiology and biotechnology , biochemistry , gene , genome , base sequence
This study provides a rapid, simple, high-throughput method to discover novel archaeal nucleic acid modifying enzymes by utilizing a fosmid genomic library, next-generation sequencing, and capillary electrophoresis. The method described here provides the details necessary to create 384-well fosmid library plates fromThermococcus kodakarensis genomic DNA, sequences of 384-well fosmids plates using Illumina next-generation sequencing, and performed high-throughput functional read-out assays using capillary electrophoresis to identify a variety of nucleic acid modifying activities, including DNA cleavage and ligation.