Sinorhizobium meliloti genome

Dr. Sharon R. Long and colleagues
Dept. of Biology at Stanford and
DNA Sequencing & Technology Center at Stanford

Sinorhizobium meliloti (formerly Rhizobium meliloti) is a soil bacterium that forms nitrogen-fixing nodules on the roots of certain genera of leguminous plants, including Medicago, Melilotus and Trigonella. Like many other members of the alpha subdivision of the proteobacteria (purple bacteria), S. meliloti possesses a multipartite genome. The S. meliloti genome contains three replicons: a 3.65-Mb chromosome and two megaplasmids, pSyma (1.35 Mb) and pSymb (1.68 Mb).


S. meliloti cells.
SEM courtesy of William Margolin.


alfalfa (Medicago sativa)

In July 1999, as part of a joint project between Dr. Sharon R. Long's group (Dept. of Biology) and the Stanford Genome Technology Center, we completed a 1X-random pass of the S. meliloti strain 1021 genome consisting of 13,617 sequence reads and representing a theoretical coverage of 63% of the genome.

This 1X-random pass provided a glimpse of the genome and allowed us to optimize protocols for the complete sequencing of the S. meliloti pSymA megaplasmid. The pSymA sequence project was part of an internation consortium to sequence the entire genome of S. meliloti.

We contructed a S. meliloti genomic library enriched for pSymA DNA. This library was constructed by purifying the 1.35-Mb pSyma DNA, which had been linearized by digestion with the restriction enzyme SwaI, via pulsed field gel electrophoresis (see here for methods). The resulting DNA was sheared, size selected for 1- to 2-kb pieces, cloned into a linker/adaptor version of M13mp18 to minimize chimera formation (R.W. Hyman et al., unpublished) and sequenced. The high-resolution physical map of S. meliloti pSymA was used to guide correct assembly of contigs.

Invasion of S. meliloti cells, overexpressing green fluorescent
protein (GFP), into an alfalfa root (red). This root hair contains a
rare, double infection thread (Gage et al. 1996).

The pSymA sequence was reported concurrently with the sequence of pSymB and the chromosome in Proceedings of the National Academy of Sciences USA. A publication that integrates the data from all three replicons was published in Science July 27, 2001 Volume 293:668-672

Links to Science publication [Abstract] [Full text]

Link to pSymA sequence publication at PNAS August 14, 2001 Volume 98:9883-9888

Link to pSymB sequence publication at PNAS August 14, 2001 Volume 98:9889-9894

Link to chromosome sequence publication at PNAS August 14, 2001 Volume 98:9877-9882

The sequence data can be accessed at the S. meliloti strain 1021 genome project website or at NCBI

 

Left panel: Mature, nitrogen-fixing nodules on an alfalfa root.
Right panel: Thin section of alfalfa nodule stained with DAPI and
acridine orange. An infection thread is visible as a white-blue
fluorescing structure in the outer layers (right side) [photo by Mark Dudley].

Selected references

Please send questions or comments to Melanie Barnett.