India-UK Nitrogen Fixation Centre
Lead Research Organisation:
John Innes Centre
Department Name: UNLISTED
Abstract
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Technical Summary
In this proposal we will use traditional microbial science as well as synthetic biology to engineer synthetic interactions between diazotrophs and cereals. This is driven by the need to improve the sustainability of Indian agriculture, namely decreasing the addition of N-fertilizer while maintaining yields.
We will utilise A. caulinodans ORS 571 as a model endophyte to engineer ammonium excretion in association with rice. Regulatory networks controlling nitrogen regulation of nif gene expression will be disrupted, firstly by removing the amino-terminal regulatory domain of the NifA activator protein and secondly by down-regulating the level of the GlnB and GlnK signal transduction proteins. The latter manipulation also influences deadenylylation of glutamine synthetase in A. caulinodans, resulting in ammonium excretion (Michel-Reydellet & Kaminski, 1999). Other key proteins involved in ammonia assimilation (e.g. GOGAT, GDH) and the ammonium transporter AmtB will also be targeted. To ensure that engineered bacteria remain competitive in the rhizosphere, we will utilise plant colonisation-specific gene switches to ensure that components required for ammonium release are only switched on or off when A. caulinodans associates with the crop. To ensure we have selected appropriate rhizosphere switches for control we will also conduct RNASeq of these strains in the rice rhizosphere.
We will utilise A. caulinodans ORS 571 as a model endophyte to engineer ammonium excretion in association with rice. Regulatory networks controlling nitrogen regulation of nif gene expression will be disrupted, firstly by removing the amino-terminal regulatory domain of the NifA activator protein and secondly by down-regulating the level of the GlnB and GlnK signal transduction proteins. The latter manipulation also influences deadenylylation of glutamine synthetase in A. caulinodans, resulting in ammonium excretion (Michel-Reydellet & Kaminski, 1999). Other key proteins involved in ammonia assimilation (e.g. GOGAT, GDH) and the ammonium transporter AmtB will also be targeted. To ensure that engineered bacteria remain competitive in the rhizosphere, we will utilise plant colonisation-specific gene switches to ensure that components required for ammonium release are only switched on or off when A. caulinodans associates with the crop. To ensure we have selected appropriate rhizosphere switches for control we will also conduct RNASeq of these strains in the rice rhizosphere.
Planned Impact
unavailable
