Working group session:
Structural Genomics
Presentation type:
poster
Authors:
Nyaku, Seloame ; Kantety, Ramesh; Sharma, Govind
Presenter:
Nyaku, Seloame
Correspondent:
Kantety, Ramesh; Sharma, Govind
Abstract:
The reniform nematode (RN) is a devastating pest of cotton and the lack of sequence data for this organism is a limitation in its management. Therefore understanding of its parasitism, virulence, and reproductive efficacy are necessary. Development of whole-genome amplification (WGA) techniques for amplification of DNA from nanogram quantities has aided the success of most genetic research. These methods prove very useful in applications such as sequencing genomes of single nematodes. Two RN libraries were generated from Whole Genome Amplification (WGA) kit (Sigma-Aldrich), and the REPLI-g kit (Qiagen). These libraries were from four female surface-sterilized RN pooled together. Prior to generation of the libraries, DNA was extracted from single RN and amplified with universal bacterial primers for verification of bacterial contamination. The DNA from the seven RN were further amplified with 18S rRNA primers, cloned, and sequenced on an ABI 3100 sequencer. The sequences were then screened for homology to nematode sequences using the standard nucleotide-nucleotide BLAST [blastn] on the NCBI website (http://www.ncbi.nlm.nih.gov). Sequences from the RN having high homology to reniform nematode sequences in the GenBank were noted and the DNA from four selected RN were used for the WGA amplifications and 454 genome sequencing. Sequence assemblies from both libraries produce combined lengths of 4 Mb and 12 Mb for both REPLIg and SIGMA libraries respectively. Initial genome characterization shows high levels of low complexity repeats in both libraries. Additional genes present in the RN genome will be characterized through comparative analysis with GenBank and other databases.