A rapid,novel and high-throughput identification of putative bell pepper transformants generated through in planta transformation approach

The recent progress on bell pepper transformation through in planta approach is incredibly simple with high transformation efficiencies in indigenous varieties. This method produces chimeric primary transformants (T₀) hence, a large number of T₀ plants and an efficient high throughput screening strategy is necessary to be developed. In the present investigation, we have standardized a rapid and highly efficient method of screening putative bell pepper transformants. Bell pepper transformants harbouring gfp genes was developed using in planta approach and 1050 etiolated T₁ seedlings were screened using fluorescent microscope to select GFP positive plants. The molecular analysis such as, gene specific PCRs, semiquantitative RT-PCR and Southern blots analysis corroborated the transgene integration in selected T₁ transformants. Based on progressive selection strategy, the transformation efficiency was 27.4% in gfp screened plants. The fluorescence based screening is viable, rapid, requires minimal labour, expense, and expertise with less percent of non-transformant escapes; further, this method can be extended from bell pepper to other crops to identify putative transformants developed through in planta approach.     

An in vivo root hair assay for determining rates of apoptotic-like programmed cell death in plants

ABSTRACT: In Arabidopsis thaliana we demonstrate that dying root hairs provide an easy and rapid in vivo model for the morphological identification of apoptotic-like programmed cell death (AL-PCD) in plants. The model described here is transferable between species, can be used to investigate rates of AL-PCD in response to various treatments and to identify modulation of AL-PCD rates in mutant/transgenic plant lines facilitating rapid screening of mutant populations in order to identify genes involved in AL-PCD regulation.     

Forest type influences transmission of Phytophthora ramorum in California oak woodlands

The transmission ecology of Phytophthora ramorum from bay laurel (Umbellularia californica) leaves was compared between mixed-evergreen and redwood forest types throughout winter and summer disease cycles in central, coastal California. In a preliminary multisite study, we found that abscission rates of infected leaves were higher at mixed-evergreen sites. In addition, final infection counts were slightly higher at mixed-evergreen sites or not significantly different than at redwood sites, in part due to competition from other foliar pathogens at redwood sites. In a subsequent, detailed study of paired sites where P. ramorum was the main foliar pathogen, summer survival of P. ramorum in bay laurel leaves was lower in mixed-evergreen forest due to lower recovery from infected attached leaves and higher abscission rates of infected leaves. Onset of inoculum production and new infections of bay laurel leaves occurred later in mixed-evergreen forest. Mean inoculum levels in rainwater and final infection counts on leaves were higher in redwood forest. Based on these two studies, lower summer survival of reservoir inoculum in bay laurel leaves in mixed-evergreen forest may result in delayed onset of both inoculum production and new infections, leading to slower disease progress in the early rainy season compared with redwood forest. Although final infection counts also will depend on other foliar pathogens and disease history, in sites where P. ramorum is the main foliar pathogen, these transmission patterns suggest higher rates of disease spread in redwood forests during rainy seasons of short or average length.     

Stability in the composition equivalence of grain from insect-protected maize and seed from glyphosate-tolerant soybean to conventional counterparts over multiple seasons, locations, and breeding germplasms

Insect-protected maize MON 810 and Roundup Ready soybean 40-3-2 represent major milestones in the adoption of genetically modified (GM) crops to enhance agricultural productivity. This study provides an assessment of the compositional stability of these products over multiple seasons, multiple germplasms, and diverse geographies encompassing North, Central, and South America and Europe. The compositional assessment evaluated levels of proximates in MON 810 and proximates, antinutrients, and isoflavones in 40-3-2. The means and range values for component levels in the GM crops and their conventional comparators were consistently similar to each other within each corresponding year from 2000 to 2009. To our knowledge, this study represents the first meta-analysis of comparative composition assessments of GM products. This approach, combined with graphical approaches, provided an effective summary of the overall data set and confirmed the continued compositional equivalence of these important crops to their conventional counterparts over time.     

Photosynthetic responses of field-grown Pinus radiata trees to artificial and aphid-induced defoliation

The phloem-feeding aphid Essigella californica represents a potential threat to the productivity of Pinus radiata plantations in south-eastern Australia. Five- and nine-year-old field trials were used to characterize the effects of artificial and natural aphid-induced (E. californica) defoliation, respectively, on shoot photosynthesis and growth. Photosynthetic capacity (A(max)) was significantly greater following a 25% (D25) (13.8 μmol m(-2) s(-1)) and a 50% (D50) (15.9 μmol m(-2) s(-1)) single-event upper-crown artificial defoliation, 3 weeks after defoliation than in undefoliated control trees (12.9 μmol m(-2) s(-1)). This response was consistently observed for up to 11 weeks after the defoliation event; by Week 16, there was no difference in A(max) between control and defoliated trees. In the D50 treatment, this increased A(max) was not sufficient to fully compensate for the foliage loss as evidenced by the reduced diameter increment (by 15%) in defoliated trees 36 weeks after defoliation. In contrast, diameter increment of trees in the D25 treatment was unaffected by defoliation. The A(max) of trees experiencing upper-crown defoliation by natural and repeated E. californica infestations varied, depending on host genotype. Despite clear differences in defoliation levels between resistant and susceptible genotypes (17 vs. 35% of tree crown defoliated, respectively), growth of susceptible genotypes was not significantly different from that of resistant genotypes. The observed increases in A(max) in the lower crown of the canopy following attack suggested that susceptible genotypes were able to partly compensate for the loss of foliage by compensatory photosynthesis. The capacity of P. radiata to regulate photosynthesis in response to natural aphid-induced defoliation provides evidence that the impact of E. californica attack on stem growth will be less than expected, at least for up to 35% defoliation.     

Purification of alkylamides from Echinacea angustifolia (DC.) Hell. Roots by high-speed countercurrent chromatography

High-speed countercurrent chromatography (HSCCC) was used for the separation of alkylamides from the roots of Echinacea angustifolia (DC.) Hell. For this purpose, the alkylamides were extracted with hexane and subjected to semipreparative HSCCC using a two-phase solvent system consisting of n-hexane, ethyl acetate, methanol, and water (4:1:2:1). The lower aqueous phase was used as the mobile phase at a flow rate of 3 mL/min and a rotary speed of 1000 rpm. This procedure led to the isolation of four pure alkylamides, that is, dodeca-2E,4E,8Z,10E/Z-tetraenoic acid isobutylamide (38.9 mg, 97% purity), dodeca-2E,4E,8Z-trienoic acid isobutylamide (4.4 mg, 92% purity), dodeca-2E,4E-dienoic acid isobutylamide (3.2 mg, 99% purity), and dodeca-2E,4E-dienoic acid 2-methylbutylamide (0.3 mg, 92% purity). The identity and purity of the isolated alkylamides were confirmed by LC-ESI-MS and (1)H NMR and (13)C NMR data. To the best of the authors' knowledge, this is the first report of dodeca-2E,4E-dienoic acid 2-methylbutylamide in E. angustifolia roots.