Optimized Replication of Arrayed Bacterial Mutant Libraries Increases Access to Biological Resources

Abstract

ABSTRACT Biological collections, including arrayed libraries of single transposon (Tn) or deletion mutants, greatly accelerate the pace of bacterial genetic research. Despite the importance of these resources, few protocols exist for the replication and distribution of these materials. Here, we describe a protocol for creating multiple replicates of an arrayed bacterial Tn library consisting of approximately 6,800 mutants in 96-well plates (73 plates). Our protocol provides multiple checkpoints to guard against contamination and minimize genetic drift caused by freeze/thaw cycles. This approach can also be scaled for arrayed culture collections of various sizes. Overall, this protocol is a valuable resource for other researchers considering the construction and distribution of arrayed culture collection resources for the benefit of the greater scientific community. IMPORTANCE Arrayed mutant collections drive robust genetic screens, but few protocols exist for replication of these resources and subsequent quality control. Increasing the distribution of arrayed biological collections will increase the accessibility and use of these resources. Developing standardized techniques for replication of these resources is essential for ensuring their quality and usefulness to the scientific community.

Keywords

• Enterococcus faecalis
• biological collections
• functional genomics
• genetic tools
• scientific rigor
• transposon mutagenesis