IDENTIFICATION OF MIRNA SEQUENCE CANDIDATE FOR DEVELOPMENT OF BIOPESTICIDE AGAINST PHYTOPHTHORA SOJAE FROM SOYBEAN (GLYCINE MAX) LINES

• Main Author: Ghildan A R, M
• Format: Final Project
• Language: Indonesia
• Subjects: Ilmu hayati ; Biologi
• Online Access: https://digilib.itb.ac.id/gdl/view/55456
• Institution: Institut Teknologi Bandung

Soybeans are important legumes because of its nutriotional content and versatility to be processed into various products. However, production of soybean is facing the problem of Phytophthora sojae attack which causes stem and root rot at a cost of up to USD 1-2 billion annually. One potential solution is by developing RNA-based biopesticides, which consist of short RNA molecules able to suppress pathogens gene expression and reduce its pathogenicity. In nature, such mechanisms are mediated by small RNAs such as miRNA. Therefore, in this study, we identify miRNA of resistant soybean lines Rps1-a, Rps1-c, and Rps1-k which could potentially be used as biopesticide candidates upon infection by P. sojae. In silico analysis conducted using smallRNA sequencing data from Zhao et. al. (2015) available on NCBI repository with accession code GSE56859. Raw reads data were quality controlled using FastQC v3.1093 followed by adapter trim using CutAdapt v3.1. Clean reads were subsequently aligned to soybean genome from the web page http://www.plantsrnas.org/ using BWA v0.7.17 and the results were quantified using HTSeq v0.11.2. The count results were filtered to eliminate too low values and normalized using the TMM (Trimmed Mean of M-value) method. Furthermore, dispersion value were constructed from housekeeping gene and differential expressions analysis is carried out using general linear model-likelihood ratio test (glmLRT) by edgeR v3.30.3. miRNA with log2-fold-change (log2FC) value > 1 were selected for target prediction analysis using FastX-toolkit v0.0.14 for reverse complement and BWA v0.7.17 for alignment against the Phytophthora sojae v3.0 (Refseq) genome. Differential expression analysis result showed that 9 specific miRNAs had increased expression of Log2FC > 1 in resistant lines and not in susceptible lines. Target prediction results showed reads from 3 of the 9 miRNAs can complement with Phytophthora sojae’s mRNA that plays a role in cell redox homeostasis, microtubule associated complex, etc.