Growth Recovery Assay and FACS-based Population Sorting Following Territorial Exclusion in Proteus mirabilis.
Bio Protoc
; 10(5): e3543, 2020 Mar 05.
Article
em En
| MEDLINE
| ID: mdl-33659517
ABSTRACT
Many bacteria take part in self recognition and kin discrimination behavior using contact-dependent effectors. Understanding the effects these effectors cause is important to explain bacterial community formation and population dynamics. Typically, kin discrimination effectors are toxins that kill target cells; their effect is therefore obvious and easily measurable. However, many self-recognition effectors, such as the Proteus mirabilis Ids system, are non-lethal and do not cause obvious physiological changes in target cells. Previously, experimental techniques to probe cells experiencing non-lethal kin recognition have been limited. Here we describe a technique to reliably isolate cells deemed self and non-self through Ids self-recognition for downstream phenotypic analysis. Liquid cultures of fluorescently labeled self-recognition mutants are mixed together and inoculated on swarm-permissive agar. Mixed swarms are harvested, and each strain is isolated through fluorescence-activated cell sorting (FACS). The growth rate of each strain is measured on a plate reader. This protocol is adaptable for other bacterial species. We describe briefly how sorted particles can be used for other analyses such as RNA-Seq library preparation.
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Coleções:
01-internacional
Base de dados:
MEDLINE
Idioma:
En
Ano de publicação:
2020
Tipo de documento:
Article