2010 Annual Report
1a.Objectives (from AD-416)
The specific objectives of this proposal are to: (1) Sequence and assemble the complete genome of P. triticina isolate 1-1, Race 1 (BBBD), which is estimated to be between 100 - 120 Mb, using a hybrid of 454 and ABI (Sanger) Fosmid-end sequence; (2) Annotate gene structure using computational methods, 200,000 454 reads of ESTs from each of four new cDNA libraries, and other available ESTs; (3) Evaluate P. triticina polymorphism and diversity by comparing the sequenced strain with three additional isolates using Illumina/Solexa sequence; (4) Prompt public release of all reads, assemblies, annotation, and discovered polymorphisms; and (5) Develop education, training and outreach programs.
1b.Approach (from AD-416)
To achieve our objective we propose to:
1. Generate a whole genome shotgun assembly using an optimized combination of traditional and new sequencing technology data (Sanger and 454).
2. Annotate gene structure using computational methods, 200,000 454 reads of ESTs from each of four new cDNA libraries, and other available ESTs.
3. Evaluate the level of P. triticina DNA polymorphisms and diversity by sequencing three additional isolates of P. triticina using Illumina/Solexa sequence.
4. Prompt public release of all reads, assemblies, annotation, discovered polymorphisms, and EST alignments.
5. Develop education, training and outreach programs
The project is to sequence the genome of Race 1 (BBBD) of Puccinia triticina Eriks (Pt). BBBD was first collected in the 1950’s in Canada and is the most avirulent race of leaf rust that has been collected, meaning most leaf rust disease resistance genes in wheat protect against this race. The Genome Sequencing Center at the Broad Institute used next generation Roche 454 sequencing and released an early assembly of the genome in January of 2010. The genome is currently estimated at ~163 Mb with 11,638 recognizable genes. The current assembly is still very fragmented, and in July, the project was granted an extension for more sequencing. An additional library of 8 kb inserts from Race 1 was made and is currently in the que for sequencing. This will allow the computer programs to reduce the fragmentation in the assembly and increase the accuracy. In addition to Race 1, three other races with varying virulence on wheat were sequenced using Illumina sequencing. The sequences from these races will be used to compare races and determine where the genomes are different.
Progress on this agreement is monitored by regularly discussing program goals and approaches and by reviewing annual accomplishments reports.