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Researcher Dash, Manoranjan
Advisor Rao, Uma
Source KrishiKosh-Indian National Agricultural Research System
Content type Text
Educational Degree Master of Science (M.Sc.)
Publisher Division of Nematology ICAR- Indian Agricultural Research Institute New Delhi
File Format PDF
Language English
Subject Domain (in DDC) Technology ♦ Agriculture & related technologies ♦ Plant injuries, diseases & pests
Subject Keyword In Silico Identification ♦ Characterization and Functional Validation of Neuropeptide-like Protein Genes (nlps) From Meloidogyne Incognita ♦ Nematology
Abstract Plant-parasitic nematodes (PPNs) account for annual global agricultural losses of an estimated $US 173 billion. Root-knot nematodes (Meloidogyne species) are the most economically important group of PPNs worldwide, with a host range that spans across most cultivated crops including vegetables and fruit crops. The potential host range of these obligate, sedentary endoparasites comprises of more than 3000 host plant species. Recent genome sequencing projects of several PPNs including Meloidogyne incognita provide an invaluable wealth of gene sequences which can be used for disturbing the nematode life cycle that in turn can help in target identification for their management options. Use of comparative genomics through bioinformatics tools can help in identification and characterization of the potential gene targets but they have to be functionally validated. RNA interference (RNAi) has emerged as a very promising tool for understanding the role of genes for their candidature in designing nematode resistant crops. In the present study, we have given transcriptional evidence for 13 Neuropeptide like protein (nlp) genes in M. incognita. The differential gene expression of two nlp genes (nlp-3 and nlp-12) in five different developmental stages revealed their higher expression in the mature females followed by pre parasitic juveniles. In the present study, functional validation using RNAi was carried out for two nlp genes, nlp-3 and nlp-12. Higher expression of these genes in the pre-parasitic stages and mature females revealed that, these genes might be involved in the host recognition and reproduction process. The in situ hybridization studies of these genes revealed their expression in neurons associated with the basal bulb and tail region of the nematodes. Both the nlp genes were silenced through RNAi which significantly reduced the number of nematodes attracted towards host at different time intervals. Silencing of nlp-12 gene was most effective in preventing the penetration up to 34.63 and 19.33 per cent at 24 and 72 hours respectively after inoculation. Further, the silencing effects on the phenotype could be correlated with the transcript abundance of the target genes by qRT-PCR. After dsRNA soaking, nlp-3 and nlp-12 were downregulated 54 and 7 folds respectively. The long term effects of in vitro RNAi was evident as it reduced the disease intensity in terms of galling in the range of 23 to 31 per cent due to reduced nematode infection. There was a 22 to 31 per cent reduction in the number of endoparasites. Additionally, it was very impressive to note that a onetime dsRNA soaking lasted till the end of nematode life cycle as reflected by reduced nematode reproduction in the range of 24 to 32 per cent and fecundity by 17 to 30 per cent. Based on the present findings, it could be concluded that RNAi of nlp- 3 and nlp-12 genes indicated them as potential gene targets for effective nematode management through host delivered RNAi for enhanced level of disease reduction.
Educational Use Research
Education Level UG and PG
Learning Resource Type Thesis
Size (in Bytes) 2.79 MB