A DE NOVO ASSEMBLY METHOD FOR SHORT SEQUENCE OF SOLID-SAGE READS RESPONSIBLE FOR WHEAT (TRITICUM AESTIVUM L.) LEAF RUST
Objective: Wheat leaf rust is one of the most widespread rust diseases caused by Puccinia triticina Eriks. De novo assembly of short sequence reads in order to understand the molecular phenomenon underlying wheat leaf rust interaction and to assemble differentially expressed genes, resistance genes and the genes encoding transcription factors in response to Puccinia infection in wheat was the main objective of the present study.
Methods: De novo assembly of SOLiD (sequencing by oligonucleotide ligation and detection) SAGE (serial analysis of gene expression) sequence reads from a pair of Near-isogenic lines (NILs) of wheat cultivar HD2329 with Lr28 (resistant) and HD2329 lacking Lr28 (susceptible) that were either infected with the most virulent pathogen Puccinia triticina or inoculated as mock in the absence of any reference sequence was carried out using multiple k-mer approach. Combinations of different software working on different algorithm were used to obtain a maximum number of differentially expressed transcripts.
Results: De novo assembly at different k-mers produced a large number of contigs. The size of contigs was further increased with the use of different assembly software. Redundancy was removed both at nucleotide and protein levels, which increased the quality of assembly.
Conclusion: For the assembly of short sequences of the complex genome such as those of polyploids a combination of software gives longer and unique contigs. It may be used in understanding the molecular mechanism of plant-microbe interaction.
Keywords: Wheat, Leaf rust, SOLiD, SAGE, De novo assembly, NILs.
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