Application of Transposon Insertion Sequencing to Agricultural Science

Abstract

Many plant-associated bacteria have the ability to positively affect plant growth and there is growing interest in utilizing such bacteria in agricultural settings to reduce reliance on pesticides and fertilizers. However, our capacity to utilize microbes in this way is currently limited due to patchy understanding of bacterial-plant interactions at a molecular level. Traditional methods of studying molecular interactions have sought to characterize the function of one gene at a time, but the slow pace of this work means the functions of the vast majority of bacterial genes remain unknown or poorly understood. New approaches to improve and speed up investigations into the functions of bacterial genes in agricultural systems will facilitate efforts to optimize microbial communities and develop microbe-based products. Techniques enabling high-throughput gene functional analysis, such as transposon insertion sequencing analyses, have great potential to be widely applied to determine key aspects of plant-bacterial interactions. Transposon insertion sequencing combines saturation transposon mutagenesis and high-throughput sequencing to simultaneously investigate the function of all the nonessential genes in a bacterial genome. This technique can be used for both in vitro and in vivo studies to identify genes involved in microbe-plant interactions, stress tolerance and pathogen virulence. The information provided by such investigations will rapidly accelerate the rate of bacterial gene functional determination and provide insights into the genes and pathways that underlie biotic interactions, metabolism, and survival of agriculturally relevant bacteria. This knowledge could be used to select the most appropriate plant growth promoting bacteria for a specific set of conditions, formulating crop inoculants, or developing crop protection products. This review provides an overview of transposon insertion sequencing, outlines how this approach has been applied to study plant-associated bacteria, and proposes new applications of these techniques for the benefit of agriculture.

Genomic Analyses Identify Manganese Homeostasis as a Driver of Group B Streptococcal Vaginal Colonization

“…Interestingly, cylE mutants have previously been shown to exhibit a decreased persistence in a murine model of vaginal colonization ( 43 ). Thus, there are limitations to the sensitivity of transposon sequencing, specifically as it is unable to detect the contribution of individual mutations in genes encoding products secreted into the environment ( 44 , 45 ). Also in this study, rather than declaring gene essentiality, we aimed to focus our analyses on GBS transposon mutant fitness.…”

A decade of advances in transposon-insertion sequencing

“…Others have identified genes in E. coli that promote survival during exposure to ionizing radiation 96 , genes in Salmonella Typhi that allow adaptation to survival in water 97 and genes involved in desiccation stress in Salmonella Typhimurium 98 . TIS approaches have also uncovered how bacteria interact with plants or deal with soil environments 99 . For instance, TIS was used to identify genes required for growth of the soft-rot pathogenic bacterium Dickeya dadantii in chicory plants 100 or genes of Pantoea stewartia that are essential for survival in planta to provide insights into how it causes wilt disease in corn 101 .…”

Genome-Wide Identification of Ralstonia solanacearum Genes Required for Survival in Tomato Plants

“…Transposon insertion sequencing (Tn-seq) is a high-throughput approach that couples genome-wide transposon mutagenesis with next-generation sequencing ( 15 , 16 ). Tn-seq can be used to identify genes that contribute to bacterial survival under the selective pressure of interest ( 17 ). Tn-seq has been applied to identify genes important for in vivo survival in several plant-pathogenic bacteria using the host environment as the selective pressure.…”