The N. benthamiana draft genome v2.6.1. is a new assembly version based on the Illumina data used in the version Niben1.0.1, with the addition of new Illumina mate pair data (20Kb, 15Kb and 5Kb) produced by Prof. Greg Martin and Prof. Lukas Mueller (BTI), PacBio Sequel Long reads produced by Dr. Brain Kivtko (UGA) and HiC data donated by Dr. Silin Zhong (HKCU). The result is an assembly with 19 pseudomolecules and 17,620 small scaffolds.

Assembly

The assembly was performed using SOAPdenovo2 with the default settings on the Illumina data (PE500bp + MP2Kb +MP5Kb + MP15Kb + MP20Kb) with one round of gap filling with GapCloser (Illumina PE500bp) and another round with PBJelly (10X coverage with PacBio Sequel). The assembly was re-scaffolded again with SSPACE-Long (PacBio Sequel) and the gaps filled again with another round of PBJelly. This final assembly was scaffolded into chromosomes using HiC data and the tool 3D-DNA.

STATS

• Total assembly size (Gb): 3.03.
• Total number of sequences: 19 chromosomes (>95% anchoring) + 17,620 scaffolds.
• Longest sequence (Mb): 194.60.
• N90 (sequences): 18
• L90 (Mb): 137.64
• BUSCO: C:95.9%[S:47.6%,D:48.3%],F:2.1%,M:2.0%
• C:97.7%[S:46.6%,D:51.1%],F:2.0%,M:0.3%,n:1375
• Merqury:
  • Completeness: 98.12%
  • QV: 29.4
  • Consensus error rate: 0.001

Annotation

The annotation was performed with Maker-P. In brief, we downloaded all the RNA-Seq publicly available for N. benthamiana from NCBI SRA (this was in 2017) and we mapped to the Niben2.6.1 reference with STAR. A evidence-based set of gene models was created with StringTie. A de-novo repeat library was generated with RepeatModeler2. Then, the models created with StringTie were used to train Augustus using Braker. The models generated with StringTie, the training files from Augustus, the protein set from different Solanaceae genomes and the repeat library from RepeatModeler2 were used as inputs for Makert-P.

• Gene models: 61,328.
• Exons/gene model: 4.98
• Annotated 5' UTRs: 1,901
• Annotated 3' UTRs: 244
• Average protein length (Aa): 333
• Gene space size (Mb): 281.
• Repeats:
  • Low complexity: 1.27%
  • DNA Transposons: 1.25%
  • LINE Transposons: 1.57%
  • SINE Transposons: 0.36%
  • LTR/Copia: 4.60%
  • LTR/Gypsy: 10.73%
  • Other LTR transposons: 0.16%
  • rRNA: 0.03%
  • tRNA: <0.01%
  • snRNA: <0.01%

If you have any questions, please contact Professor Aureliano Bombarely.

A new draft sequence of the Nicotiana benthamiana genome has been released by researchers from the Boyce Thompson Institute for Plant Research (BTI). It is accessible through the SGN BLAST tool and JBrowse, and can be downloaded from the SGN ftp site. More information about this project can be found at the BTI Nicotiana benthamiana Project site

N. benthamiana has a very complex genome consisting of n=19 chromosomes that is estimated at over 3.5GB in size. Taxonomically, it is an old allotetraploid of the section Suaveolentes that was formed from diploid parents from the section Sylvestres and the section Noctiflorae. It is native to Australia [2, 3].

N. benthamiana is a widely used model for plant-microbe biology and other research applications. It is particularly useful because it is related to tomato and potato and is amenable to virus-induced gene silencing (VIGS) which facilitates the efficient functional study of plant genes.

The draft sequence of N. benthamiana will allow better design of constructs for virus-induced gene-silencing (VIGS) in order to reduce the possibility of 'off-target' gene silencing [1]. The sequence will be also useful to identify tomato and potato orthologs for comparative genomics with other Solanaceae species and, to retrieve promoter sequences.

Data Agreement

Accessing to the Nicotiana benthamiana genome data, you agree not to publish any article containing any analysis of the whole gene space or whole genome scale prior to the publication by the Benthamiana Sequencing and Annotation Consortium (BSAC). These analysis include the identification of complete (whole genome) sets of genomic features such as genes, gene families, regulatory elements, repeat structures, GC content, or any other genome feature, and whole-genome- or chromosome- scale comparisons with other species. Scientific users are free to publish analysis based in a single gene (or group of genes).

If you have any questions, please contact Professor Gregory Martin or Professor Aureliano Bombarely.

Assembly Statistics

Sequencing approach

Seedlings were grown in the greenhouse and nuclear DNA was extracted from etiolated leaves. From the DNA, three distinct libraries were synthesized for use in a Illumina HiSeq-2000 sequencer: 1) a paired end library with an insert size of approximately 500 bp insert size; 2) a mate-pair type library with an insert size of 2kb; and 3) a mate pair with an insert size of 5kb. Sequencing was performed using a full run with 4 lanes of the paired end library and 1 lane each of the mate pair libraries.

Accession sequenced

The accession sequenced is referred to as "Nb-1". Seeds can be obtained by contacting Professor Gregory Martin.

Links

• More information about this project at the BTI Nicotiana benthamiana Project site
• Austrialia/New Zealand Nicotiana benthamiana sequencing project
• On-line tutorial for VIGS procedure

References

1. Bombarely, A., H. G. Rosli, J. Vrebalov, P. Moffett, L. A. Mueller, and G. B. Martin (2012). A draft genome sequence of Nicotiana benthamiana to enhance molecular plant-microbe biology research. Molecular Plant-Microbe Interactions 25:1523-1530.
2. Knapp, S. M. W. Chase and J. J. Clarkson (2004). Nomenclatural changes and a new sectional classification in Nicotiana (Solanaceae). Taxon 53:73-82.
3. Goodin, M. M., D. Zaitlin, R. A. Naidu, and S. A. Lommel (2008). Nicotiana benthamiana: Its history and future as a model for plant-pathogen interactions. Molecular Plant-Microbe Interactions 21:1026.

Acknowledgments

Sequencing of the N. benthamiana genome was supported by funds from the Boyce Thompson Institute for Plant Research and National Science Foundation grants IOS-1025642 (A. Collmer and G. Martin), and IOS-0343327 (P. Moffett).