Anil Kumar Mishra

Silencing potential of viral derived RNAi constructs in Tomato leaf curl virus -AC4 gene suppression in tomato

We investigated viral gene suppression in an infected tomato, by transforming it with RNA inhibition (RNAi) constructs derived from same viral gene. To develop RNAi constructs, conserved sequences ranging from 21 to 200 nt of the viral target AC4 gene of various viruses causing the tomato leaf curl disease were chosen. The double-stranded (ds)RNA producing constructs carry the sense and antisense portions of these sequences and are separated by different introns behind a constitutive promoter. We compared the levels of suppression of the viral target gene by transforming four different RNAi constructs with varied arm length of dsRNA. Gene silencing levels of the viral target gene were found to be directly proportional to the arm length of the dsRNA. We observed that dsRNA derived from longer arm-length constructs generating a pool of siRNAs that were more effective in targeting gene silencing. After transformation, one of the RNAi construct having a 21 nt arm-length produced aberrant phenotypes. These phenotypic anomalies may be due to unintended (‘off-target’) host transcript silencing. The unintended host transcript silencing showed modest reversion in the presence of the viral target gene. The findings presented here suggest that the arm length of dsRNA capable of producing a pool of diced siRNAs is more efficient in gene silencing, the effect of off-targeting siRNA is minimized in a pool, and off-targeting silencing can be minimized in the presence of target gene.

Comparative evaluation of horizontal accuracy of elevations of selected ground control points from ASTER and SRTM DEM with respect to CARTOSAT-1 DEM: a case study of Shahjahanpur district, Uttar Pradesh, India

Digital elevation model (DEM) data of Shuttle Radar Topography Mission (SRTM) are distributed at a horizontal resolution of 90 m (30 m only for US) for the world, Advanced Spaceborne Thermal Emission and Reflection Radiometer (ASTER) DEM data provide 30 m horizontal resolution, while CARTOSAT-1 (IRS-P5) gives 2.6 m horizontal resolution for global coverage. SRTM and ASTER data are available freely but 2.6 m CARTOSAT-1 data are costly. Hence, through this study, we found out a horizontal accuracy for selected ground control points (GCPs) from SRTM and ASTER with respect to CARTOSAT-1 DEM to implement this result (observed from horizontal accuracy) for those areas where the 2.6-m horizontal resolution data are not available. In addition to this, the present study helps in providing a benchmark against which the future DEM products (with horizontal resolution less than CARTOSAT-1) with respect to CARTOSAT-1 DEM can be evaluated. The original SRTM image contained voids that were represented digitally as −140; such voids were initially filled using the measured values of elevation for obtaining accurate DEM. Horizontal accuracy analysis between SRTM- and ASTER-derived DEMs with respect to CARTOSAT-1 (IRS-P5) DEM allowed a qualitative assessment of the horizontal component of the error, and the appropriable statistical measures were used to estimate their horizontal accuracies. The horizontal accuracy for ASTER and SRTM DEM with respect to CARTOSAT-1 were evaluated using the root mean square error (RMSE) and relative root mean square error (R-RMSE). The results from this study revealed that the average RMSE of 20 selected GCPs was 2.17 for SRTM and 2.817 for ASTER, which are also validated using R-RMSE test which proves that SRTM data have good horizontal accuracy than ASTER with respect to CARTOSAT-1 because the average R-RMSE of 20 GCPs was 3.7 × 10−4 and 5.3 × 10−4 for SRTM and ASTER, respectively.

Viral suppression in transgenic plants expressing chimeric transgene from tomato leaf curl virus and cucumber mosaic virus

The genes CMV-cp and ToLCV-cons-rep from the isolates of cucumber mosaic virus (CMV) and tomato leaf curl virus (ToLCV) were transcriptionally fused under the control of CaMV 35 S promotor. This construct was used to transform tobacco and tomato using Agrobacterium. We show transforming ToLCV and CMV infected plants with the homologous chimeric gene construct, that produces RNAs, capable of duplex formation, confers gene silencing. We proposed that the antisense suppression in ToLCV infected plants provides a threshold level of dsRNA needed to induce gene silencing where as sense suppression in CMV infected plants may be operating through cosuppression, leading to delayed and attenuated symptoms.

Behavior of RNAi suppressor protein 2b of Cucumber mosaic virus in planta in presence and absence of virus.

The 2b protein encoded by Cucumber mosaic virus (CMV) has been shown as a virus counter defense factor that interferes with the RNAi pathway. The 2b gene from CMV-banana, New Delhi isolate (CMV-NDLS) was amplified from CMV infected cucumber plants to generate the sense and antisense binary vector constructs for 2b expression and repression in planta. Constitutive expression of 2b gene in healthy Nicotiana tabacum caused phenotypic aberrations during somatic embryogenesis, which were not observed when expressed in CMV infected N. tabacum. Further, the established virus population in CMV infected N. tabacum was not affected by constitutive expression and repression of 2b gene. Thus, indicating its role in initiation of gene silencing, at the early stage of viral infection. This is the first demonstration of differential behavior of 2b suppressor protein in host development in the absence and presence of virus.

Evaluation of relief aspects morphometric parameters derived from different sources of DEMs and its effects over time of concentration of runoff (TC)

An attempt has been made to study the relief aspects from three different sources of Digital Elevation Models (DEMs) viz., Survey of India (SOI) topographic map (1:25,000), Shuttle Radar Topography Mission (SRTM-90 m and SRTM-30 m) and Advanced Spaceborne Thermal Emission and Reflection Radiometer (ASTER-30 m). These aspects are evaluated to examine differences among them and their influence on time of concentration (TC) of runoff at Moolbari Experimental Watershed (MEW), Sub Himalayan region (Shimla, District of Himachal Pradesh) India. For detailed study of relief aspects, morphometry parameters, SOI topographic map (as base map) were used. The results show that the relief aspects morphometric parameters derived from the SRTM30m and ASTER30m lie between SOI topographic map, and SRTM90m. We estimated TC of 21 micro watersheds from different sources DEMs by using Kirpich, Johnstone, Témez, and Barnsby equation only. Témez and Barnsby equation demonstrate high potential for the identification of TC from SOI Topographic map, SRTM90m, ASTER and SRTM30m DEMs. TC_Topo has a positive relationship with TC_SRTM90m, TC_ASTER and TC_SRTM30m for both Témez, and Barnsby equation with R2=0.804Topo & SRTM90m, 0.810Topo & ASTER & 0.839Topo & SRTM30m and 0.712Topo & SRTM90m, 0.747 Topo & ASTER & 0.785 Topo & SRTM30m. Further statistical test of Témez, and Barnsby equation based TC, only Témez equation based TC qualify/satisfy the statistical test. by considering all freeware DEMs a Semi-empirical model (SEM) has been developed, where TC predicted in term of TC_Topo is a function of TC_SRTM90m, TC_ASTER and TC_SRTM30m. This SEM has R2=0.883 and adjusted R= 0.874, Multiple R=0.907 and with Standard Error =2.131 at 95% confidence level. Comparison of the TC derived from the multiple regressions among three DEMs with TC_Topo shows an RMSE of 3.803, R-RMSE of 0.169, NRMSE of 0.342, R2 of 0.89, and RMSE% of 3.296 for Témez equation.

Soil suitability analysis for crop planning in kheragarah Tehsil of Agra district, Uttar Pradesh

Soil suitability is prerequisite for sustainable agriculture and it plays a pivotal role in crop production especially in degraded lands. In this study, an attempt was made to assess the soil suitability of Kheragarah tehsil for production of major crops during rabi and kharif seasons. Six pedons of the tehsil were studied for their physical and chemical properties and their suitability for growing wheat, maize, cotton and barley. All pedons were found to be moderate to deep in soil depth, well to poorly drained, slightly to strongly alkaline (pH 7.98 - 9.43), slightly to highly saline (ECe 1.81- 7.23 dS m−1), low in organic carbon (1.1–2.7 g kg−1), CEC medium to high [19.7 - 24.4 cmol(p+) kg−1]. The calcium carbonate content ranged from nil to 14.3%. Soils were low in available N (72.0 to 220.0 kg ha−1), low to high in available P (3.7–15.0 kg ha−1) and medium to high in available K (153.0 - 361.6 kg ha−1). However, the soils were adequate in available Fe and Mn but low in available Zn and Cu content. The soil suitability analysis revealed that soils at P1 locations were moderately suitable for growing wheat and barley and marginally suitable for cotton and maize cultivations. The P2 soils were moderately suitable for wheat, maize and barley and marginally suitable for cotton. However, the soils of P3 and P5 were not suitable for growing any of the selected crops. But, P6 soils were moderately suitable for growing all selected crops whereas, P4 soils were highly suitable for all selected crops except maize.

Soil Erosion Risk Assessment and Spatial Mapping in Jhagrabaria Watershed, Allahabad, U.P. (India) by Using LANDSAT 7ETM+ Remote Sensing Data, Revised Universal Soil Loss Equation (RUSLE) and Geographical Information System (GIS)

This article discusses the application of the Revised Universal Soil Loss Equation (RUSLE) in conjunction with LANDSAT 7ETM+ remote sensing data, and geographical information system (GIS) to the spatial mapping of soil erosion risk in Jhagrabaria watershed Allahabad, U.P., India. Soil map and topographical data were used to develop the soil erodibility factor (K) and a digital elevation model (DEM) image was used to generate the topographic factor (LS). The cover-management factor (C) was developed based on vegetation, shade, and soil fraction images derived from spectral mixture analysis of a LANDSAT Enhanced Thematic Mapper Plus (LANDSAT 7ETM+) image. Support practice factors (P) was developed by crossing operation between land use/land cover classification map and slope map. Assuming the same climatic conditions in the study area, the rainfall-runoff erosivity (R) factor was not used. The value of K for study area lies between 0.25 and 0.485, LS values were less than 1.4, C and P values were less than 1. A soil erosion risk map with five classes (very low, low, medium, medium-high, and high) was produced based on the simplified RUSLE within the GIS environment and was linked to land use/land cover (LULC) image to explore relationships between soil erosion risk and LULC distribution. The results indicate that most succession and mature vegetation are in low erosion risk areas, while Barren and Fallow lands are usually associated with medium to high erosion risk areas. This research implies that remote sensing and GIS provide promising tools for evaluating and mapping soil erosion risk in the Jhagrabaria watershed of India.

Differential role of tomato Gemini virus and cucumovirus in plant organogenesis.

Phenotypic symptom development in plants due to viral infection is believed to be a result of developmental defects caused by regulation of gene expression by viral suppressors. Since viral suppressors of viruses differentially regulate small RNAs (siRNA and miRNA), viral pathogenicity and symptom development in the host cannot apply universally. In present study, we demonstrated the differential effects of presence of two viruses (Tomato leaf curl virus and Cucumber mosaic virus) in organogenesis of two separate hosts (tomato and tobacco). Presence of ToLCV in the tomato explants severely affected the tomato organogenesis leading to sharp decrease in the survival percentage and severe developmental defects. On the other hand presence of CMV in the tobacco explants had not shown much effect in tobacco organogenesis as well as survival percentage. This study has not only highlighted our basic understanding of host-virus interactions in terms of viral suppressors and symptom development, but also provide an alternative way of maintaining and reviving viral cultures in vitro.