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Enhanced methanol production in plants provides broad spectrum insect resistance.

Dixit S, Upadhyay SK, Singh H, Sidhu OP, Verma PC, K C - PLoS ONE (2013)

Bottom Line: In-planta bioassay on transgenic lines showed up to 99 and 75% reduction in the population multiplication of plant sap sucking pests Myzus persicae (aphid) and Bemisia tabaci (whitefly), respectively.Confocal microscopy showed no deformities in cellular integrity, structure and density of stomata and trichomes of transgenic plants compared to WT.Cell wall enzyme transcript levels were comparable with WT.

View Article: PubMed Central - PubMed

Affiliation: CSIR-National Botanical Research Institute, Council of Scientific and Industrial Research, Lucknow, Uttar Pradesh, India ; Academy of Scientific and Innovative Research (AcSIR), Anusandhan Bhawan, 2-Rafi Marg, New Delhi, India.

ABSTRACT
Plants naturally emit methanol as volatile organic compound. Methanol is toxic to insect pests; but the quantity produced by most of the plants is not enough to protect them against invading insect pests. In the present study, we demonstrated that the over-expression of pectin methylesterase, derived from Arabidopsis thaliana and Aspergillus niger, in transgenic tobacco plants enhances methanol production and resistance to polyphagous insect pests. Methanol content in the leaves of transgenic plants was measured using proton nuclear spectroscopy (1H NMR) and spectra showed up to 16 fold higher methanol as compared to control wild type (WT) plants. A maximum of 100 and 85% mortality in chewing insects Helicoverpa armigera and Spodoptera litura larvae was observed, respectively when fed on transgenic plants leaves. The surviving larvae showed less feeding, severe growth retardation and could not develop into pupae. In-planta bioassay on transgenic lines showed up to 99 and 75% reduction in the population multiplication of plant sap sucking pests Myzus persicae (aphid) and Bemisia tabaci (whitefly), respectively. Most of the phenotypic characters of transgenic plants were similar to WT plants. Confocal microscopy showed no deformities in cellular integrity, structure and density of stomata and trichomes of transgenic plants compared to WT. Pollen germination and tube formation was also not affected in transgenic plants. Cell wall enzyme transcript levels were comparable with WT. This study demonstrated for the first time that methanol emission can be utilized for imparting broad range insect resistance in plants.

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Confocal microscopic images of transgenic and control plants.(A) Propidium iodide stained transverse section of transgenic and control plants showing well integrated cell network. (B) Structure of stomata, circumference of stomata of transgenic plant (113.48±6.7) was similar to WT (114.96±4.6). (C) Structure of trichomes (1) and stomatal density of transgenic and WT plants (2) were also similar to control plant. (D) Pollen germination of transgenic and WT (after 3 hrs). (E) FDA stained single pollen showing equal pollen tube length for transgenic and WT (1). FDA stained (2) DIC and (3) merge of FDA and DIC.
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pone-0079664-g007: Confocal microscopic images of transgenic and control plants.(A) Propidium iodide stained transverse section of transgenic and control plants showing well integrated cell network. (B) Structure of stomata, circumference of stomata of transgenic plant (113.48±6.7) was similar to WT (114.96±4.6). (C) Structure of trichomes (1) and stomatal density of transgenic and WT plants (2) were also similar to control plant. (D) Pollen germination of transgenic and WT (after 3 hrs). (E) FDA stained single pollen showing equal pollen tube length for transgenic and WT (1). FDA stained (2) DIC and (3) merge of FDA and DIC.

Mentions: PME plays vital role in plant development hence we have critically examined growth, structural and physiological parameters of transgenic plants. Transgenic plants showed normal growth pattern and morphology as compared to WT plants (Figure S2). Transgenic plants showed well integrated hexagonal cell network as observed in WT plants without any deformities (Figure 7A). Further, we examined structure and density of stomata which was also found similar in transgenic and WT plants (Figure 7B). Number of stomata on lower surface of transgenic plants (95 ± 12 per mm2) was almost similar to WT (103 ± 9 per mm2). Average circumference of stomata of transgenic plant (113.48 ± 6.7 µm) was also similar to WT (114.96 ± 4.6 µm). Naturally tobacco plants had uniseriate type of trichomes, both transgenic and WT plants showed well organized uniseriate trichomes with well arranged glandular hair (Figure 7C). PME also played critical role in pollen germination and pollen tube formation. We studied pollen viability by pollen germination assay followed by fluorescein diacetate (FDA) staining. In vitro pollen germination assay on artificial liquid media showed more than 95% pollen germination for both transgenic and WT plants (Figure 7D). Confocal microscopic images showed almost equal length of pollen tube of transgenic (3.8 ± 0.08 µm) and WT pollens (3.9 ± 0.05 µm) after 4 hrs (Figure 7E).


Enhanced methanol production in plants provides broad spectrum insect resistance.

Dixit S, Upadhyay SK, Singh H, Sidhu OP, Verma PC, K C - PLoS ONE (2013)

Confocal microscopic images of transgenic and control plants.(A) Propidium iodide stained transverse section of transgenic and control plants showing well integrated cell network. (B) Structure of stomata, circumference of stomata of transgenic plant (113.48±6.7) was similar to WT (114.96±4.6). (C) Structure of trichomes (1) and stomatal density of transgenic and WT plants (2) were also similar to control plant. (D) Pollen germination of transgenic and WT (after 3 hrs). (E) FDA stained single pollen showing equal pollen tube length for transgenic and WT (1). FDA stained (2) DIC and (3) merge of FDA and DIC.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3818224&req=5

pone-0079664-g007: Confocal microscopic images of transgenic and control plants.(A) Propidium iodide stained transverse section of transgenic and control plants showing well integrated cell network. (B) Structure of stomata, circumference of stomata of transgenic plant (113.48±6.7) was similar to WT (114.96±4.6). (C) Structure of trichomes (1) and stomatal density of transgenic and WT plants (2) were also similar to control plant. (D) Pollen germination of transgenic and WT (after 3 hrs). (E) FDA stained single pollen showing equal pollen tube length for transgenic and WT (1). FDA stained (2) DIC and (3) merge of FDA and DIC.
Mentions: PME plays vital role in plant development hence we have critically examined growth, structural and physiological parameters of transgenic plants. Transgenic plants showed normal growth pattern and morphology as compared to WT plants (Figure S2). Transgenic plants showed well integrated hexagonal cell network as observed in WT plants without any deformities (Figure 7A). Further, we examined structure and density of stomata which was also found similar in transgenic and WT plants (Figure 7B). Number of stomata on lower surface of transgenic plants (95 ± 12 per mm2) was almost similar to WT (103 ± 9 per mm2). Average circumference of stomata of transgenic plant (113.48 ± 6.7 µm) was also similar to WT (114.96 ± 4.6 µm). Naturally tobacco plants had uniseriate type of trichomes, both transgenic and WT plants showed well organized uniseriate trichomes with well arranged glandular hair (Figure 7C). PME also played critical role in pollen germination and pollen tube formation. We studied pollen viability by pollen germination assay followed by fluorescein diacetate (FDA) staining. In vitro pollen germination assay on artificial liquid media showed more than 95% pollen germination for both transgenic and WT plants (Figure 7D). Confocal microscopic images showed almost equal length of pollen tube of transgenic (3.8 ± 0.08 µm) and WT pollens (3.9 ± 0.05 µm) after 4 hrs (Figure 7E).

Bottom Line: In-planta bioassay on transgenic lines showed up to 99 and 75% reduction in the population multiplication of plant sap sucking pests Myzus persicae (aphid) and Bemisia tabaci (whitefly), respectively.Confocal microscopy showed no deformities in cellular integrity, structure and density of stomata and trichomes of transgenic plants compared to WT.Cell wall enzyme transcript levels were comparable with WT.

View Article: PubMed Central - PubMed

Affiliation: CSIR-National Botanical Research Institute, Council of Scientific and Industrial Research, Lucknow, Uttar Pradesh, India ; Academy of Scientific and Innovative Research (AcSIR), Anusandhan Bhawan, 2-Rafi Marg, New Delhi, India.

ABSTRACT
Plants naturally emit methanol as volatile organic compound. Methanol is toxic to insect pests; but the quantity produced by most of the plants is not enough to protect them against invading insect pests. In the present study, we demonstrated that the over-expression of pectin methylesterase, derived from Arabidopsis thaliana and Aspergillus niger, in transgenic tobacco plants enhances methanol production and resistance to polyphagous insect pests. Methanol content in the leaves of transgenic plants was measured using proton nuclear spectroscopy (1H NMR) and spectra showed up to 16 fold higher methanol as compared to control wild type (WT) plants. A maximum of 100 and 85% mortality in chewing insects Helicoverpa armigera and Spodoptera litura larvae was observed, respectively when fed on transgenic plants leaves. The surviving larvae showed less feeding, severe growth retardation and could not develop into pupae. In-planta bioassay on transgenic lines showed up to 99 and 75% reduction in the population multiplication of plant sap sucking pests Myzus persicae (aphid) and Bemisia tabaci (whitefly), respectively. Most of the phenotypic characters of transgenic plants were similar to WT plants. Confocal microscopy showed no deformities in cellular integrity, structure and density of stomata and trichomes of transgenic plants compared to WT. Pollen germination and tube formation was also not affected in transgenic plants. Cell wall enzyme transcript levels were comparable with WT. This study demonstrated for the first time that methanol emission can be utilized for imparting broad range insect resistance in plants.

Show MeSH
Related in: MedlinePlus