Cancelled: Biology seminar series, Jason Kearsley
Cancelled:
This week we will have an exit seminar from Jason Kearsley (Finan lab)!
There will be coffee and snacks before the seminar. Please bring your own mug.
Thursday 4:00PM, HSC 1A5 and on zoom (Passcode: cElegans)
Exploring the extrachromosomal Sinorhizobium meliloti symbiotic N2-fixing genome through extensive minimization
It is of significant agricultural and environmental interest to understand the bacterial genes and their molecular functions that are involved in the formation of N2-fixing symbioses. One method of approaching this understanding is to establish a minimal genome that is still capable of forming an effective symbiotic relationship with its host. Sinorhizobium meliloti has continuously served as an important model organism for studying the rhizobia-legume symbiosis. Most genes with direct functions in symbiotic nitrogen fixation (SNF) are harbored on two megaplasmids: pSymA (1354 kb) and pSymB (1683 kb). We have minimized the pSymA gene set to a mere 58 genes (63 kb) that were sufficient for a robust SNF phenotype. The pSymB megaplasmid is evolutionarily older and more chromosomal-like (i.e. a chromid). Several methods of minimizing the pSymB genome with respect to SNF have been conducted, including large-scale deletion analyses (top-down) and assembly-based methods (bottom-up). While pSymB has proved challenging to minimize without incurring a large symbiotic penalty, a cumulative deletion strategy has generated a strain with only 19% of pSymB (322 kb) that can still form an effective symbiosis with Medicago sativa (alfalfa). Top-down deletion approaches are useful for broad minimization, but their laborious requirements limit fine-tuning. A targeted PCR amplification of known and suspected SNF genes coupled with their assembly into distinct regions in yeast facilitated their subsequent integration into a strain lacking pSymB entirely. The 322 kb gene set was ultimately reduced to 120 genes (138 kb) while still consistently forming a functional symbiosis. The methodology employed allows easy integration of subsequent regions into this background strain to test their ability to restore SNF levels. This provides a unique way to investigate new genes of symbiotic relevance.
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