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Authors: Sanju, Suman
Title: Host-delivered RNAi-mediated silencing of Phytophthora infestans for imparting late blight resistance in potato
Language: en
Type: Thesis
Pages: 225p.
Agrotags: null
Abstract: Potato (Solanum tuberosum L.) is currently, the third most consumed crop worldwide after wheat and rice. Consumption of potato is expanding strongly in the developing nations which account for more than half of the global harvest. Late blight of potato caused by the oomycetes Phytophthora infestans, that destroys leaves, stems, and tubers, is the decimator of potato cultivation costing over $ 13 billion annually all over the world in crop losses and control measures. The strategies for managing potato late blight often remain unsustainable and costly. The control measures for late blight mainly rely on fungicides. In the present scenario, late blight resistant breeding still pose a great challenge to potato breeders, due to the breakdown of most of the resistant varieties developed earlier. RNAi is interesting mechanistically for reasons including its action at the posttranscriptional level, its exquisite sequence selectivity and the flexibility it affords for regulating gene expression. Silencing of the Avr3a gene through HD-RNAi in the transgenic background could lead to pathogen death and/or loss of virulence, and consequently imparting late blight resistance. In the detached leaf assay, among the siRNA mediated silenced lines of Kufri Khyati, least lesion area was recorded in the line K. Khyati 1129 (4.89 cm2) and K. Khyati 1037 (4.97 cm2) in comparison to other lines and control whereas in Kufri Pukhraj lines, it varied from 8.05 cm2 to 11.88 cm2 while in control it was 13.54 cm2. LER values as well as spore count was also reduced in siRNA transformed lines as compared to control plants. Whole plant bioassays against P. infestans revealed delayed and moderate resistance at 120 hpi, with a resistant nature observed at 72 hpi in four lines. Massive pathogen sporulation was observed on the wild-type in comparison to the reduced sporulation observed on susceptible transgenic lines at 120 hpi. No sporulation was observed in the moderate resistant lines of both cultivars. Transcript levels of Avr3a were significantly increased (line 2086, 4.2-fold; line 2155, 10.4-fold; line 2173, 7-fold) in leaf samples withdrawn at 0 hpi, and subsequent expression levels decreased to less than 1-fold from 72 to 120 hpi in all selected partial resistant lines compared with nontransformed controls. Pathogen DNA quantity increased progressively as the disease symptoms increased sharply over time up to 120 hpi. The quantification assay also revealed that partial resistant lines had less pathogen biomass compared with the nontransformed controls. All plants showed low intensity siRNA expression before inoculation, the expression was greatly increased in the transgenic lines (Kufri Pukhraj 2086 and 2173) compared with the control and susceptible transgenic lines not showing siRNA expression. Southern blot analysis confirmed at least one T-DNA copy integrated into the genome of the transgenic plants (Kufri Khyati 1037, Kufri Pukhraj 2173, single copy; Kufri Pukhraj 2086, three copies; Kufri Pukhraj 2155, single copy); no integration of the gene was observed in the control plants.
Description: Ph.d thesis
Subject: Botany
Theme: Host-delivered RNAi-mediated silencing of Phytophthora infestans for imparting late blight resistance in potato
Research Problem: Host-delivered RNAi-mediated silencing of Phytophthora infestans for imparting late blight resistance in potato
These Type: Ph.D
Issue Date: 2017
Appears in Collections:Thesis

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