Use Of De Novo Assembly On Illumina Transcriptome Sequencing Of Arachis Hypogaea To Distinguish Gene Isoforms.

Cultivated peanut (Arachis hypogaea L.) is an allotetraploid species whose ancestral genomes are most likely, derived from the A-genome species, A. duranensis, and the B-genome species, A. ipaensis. Rapid advances in sequencing technologies have made RNA-Seq a powerful method of choice for crops in downstream analysis. Mapping these sequences to nearly related species or ESTs can lead to loss of genome-specific information in ployploids or tissue specific information. Also, mapping to a poor reference can mask the differences of homologous and homoeologous variants. In our study we focused on generating a de novo assembly in the tetraploid peanut species using Trinity, a de novo assembler. Leaf, root and pod (yellow, brown and black maturing stage) samples from 9 different plants of spanish peanut cultivar OLin were used for total RNA isolation, and sequencing was performed on an Illumina GAIIx Analyzer at NCGR. Thirty-eight million reads obtained from RNAseq were assembled into 67,098 contigs and 30,673 were unique contigs. Assembly was performed with the default kmer of 25 and the minimum read coverage of 2. On comparing isoforms reported by Trinity along with the diploid progenitors, Trinity revealed the homoeologous nature of genes originating from two different A and B genomes. Thus, the short reads assembler was efficient in separating out homoeologs and paralogous genes. This newer transcriptome assembly will be utilized to select the homologous variants between parents of breeding populations. These variants will be validated in both tetraploid and diploid accessions.­­