Genome Assembly of 7DL

Bread wheat (Triticum aestivum L.) is one of the most important crops worldwide. The availability of its complete genome can contribute to meeting the challenges of increasing food demand and the changing global climate in the 21st century. However, wheat is an allohexaploid species (2n = 6x =42) with the genome size of 17 × 10⁹bp/1C and 90% repetitive sequences, which makes it a daunting task to sequence and completely assemble its whole genome. The International Wheat Genome Sequencing Consortium (IWGSC) has initiated to map, sequence and assemble the wheat genome based on chromosome-by-chromosome approach.

As part of the efforts of IWGSC, we constructed a physical map for wheat (Chinese Spring) chromosome arm 7DL through fingerprinting 7DL-specific BAC clones by the high information content fingerprinting technique. A total of 50,304 clones (representing ca. 15x equivalent of the chromosome arm) were fingerprinted, of which 32,765 clone-fingerprints were suitable for assembly. An automated assembly with the FPC software resulted in 1,614 contigs with 6,125 remained as singletons. The size of contigs N50 was 348kb; while L50 310 contigs. A total of 4,457 clones for Minimum Tiling Path (MTP) were established during the physical map construction. Using the Aegilops tauschiiphysical map as a template, 846 contigs of wheat 7DL chromosome were anchored and ordered.

Furthermore, the 4,457 MTP clones were sequenced individually with multiple insert size sequencing libraries using the Illumina Hiseq2000 sequencing platform. A total of about 50Gb data were generated. After de novo assembly of each clone, gap filling and the integration of the physical map, we obtained a final assembly of 1454 scaffolds representing 373 Mb, with the N50 of 391kb and N90 of 108kb, respectively. Approximately 80% of the chromosome arm sequence was covered. In addition, 3000 singltons were sequenced to fill the uncovered region. All these scaffolds are further ordered along the chromosome using the available genetic markers, such as 90K and 400K SNP arrays. Gene annotation and comparative analysis are ongoing. The ordering of contigs as well as gap-filling between contigs were greatly improved using the BAC-ends generated through our sequencing approach, demonstrating its cost-effective nature. Single molecule sequencing (Pac-Bio) was found to be quite effective in facilitating better contigs construction derived from NGS.

The physical map and reference sequences of 7DL will provide detail and ultimate information for studies on the structure, organization and evolution of this wheat chromosome, and also accelerate the identification of functional genes controlling important agronomic traits.

Key Words: wheat; 7DL; physical mapping; genome