Rapid, reliable and meaningful estimates of leaf area index (LAI) are essential to functional characterization of forest ecosystems including biomass and primary productivity studies. Accurate LAI estimates of tropical deciduous forest are required in studies of regional and global change modeling. Tropical deciduous forest due to higher species richness, multiple species association, varied phenophases, irregular stem densities and basal cover, multistoried canopy architecture and different micro-climatic conditions offers dynamism to the understanding of the LAI dynamics of different PFTs in an ecosystem. This investigation reports a new indirect method for measurement of leaf area index (LAI) in a topical moist deciduous forest in Himalayan foothills using LAI-2000 Plant Canopy Analyzer. We measured the LAI in two seasons (summer; leaf senescence stage and post-monsoon; full green stage) in three (dry miscellaneous, sal mixed and teak plantations) plant functional types (PFT) in Katerniaghat Wildlife Sanctuary, India. Ground LAI values ranged between 2.41 and 6.89, 1.17 and 7.71, and 1.92 and 5.19 during post-monsoon season and 1.36-4.49, 0.67-3.1 and 0.37-1.83 during summer season in dry miscellaneous, sal mixed and teak plantation, respectively. We observed strong correlation between LAI and community structural parameters (tree density, basal cover and species richness), with maximum with annual litter fall (R-2 > 0.8) and aboveground biomass (AGB) (R-2 > 0.75). We provided equations relating LAI with AGB, which can be utilized in future studies for this region and can be reasonably extrapolated to other regions with suitable statistical extrapolations. However, the relations between LAI and other parameters can be further improved with incorporation of data from optimized and seasonal sampling. Our indirect method of LAI estimation using litter fall as a proxy, offers repetitive potential for LAI estimate in other PFTs with relatively time and cost-effective way, thereby generating quicker and Tellable data for model run for regional and global change studies. (C) 2015 Elsevier Ltd. All rights reserved.

During November 2007, brinjal little leaf (BLL) symptoms (Fig. 1) were observed in approximately 20% of the brinjal (Solanum melongena) plants growing in the fields of Bihar, India, leading to the suspicion of a phytoplasma infection. To test for the presence of phytoplasma, genomic DNA was isolated from the leaf midribs of ten plants with and four plants without symptoms, and the phytoplasma DNA amplified by nested PCR with the universal primers P1/P7 (Deng & Hiruki, 1991) followed by R16mF2/R16mR1 (Gundersen & Lee, 1996), as previously described (Khan et al., 2004). The nested PCR amplicons of 1.4 kb corresponding to the phytoplasma 16S ribosomal DNA were cloned into pDRIVE vector (Qiagen GmbH, Germany). No PCR amplicons were observed for the symptomless plants. Twelve positive clones containing 16S ribosomal DNA of phytoplasma were sequenced and found sharing a 99.93% of sequence identity. Sequences of two clones were deposited in GenBank (Accession Nos. JQ518317 and JQ518318). In silico RFLP patterns were generated from the phytoplasma 16S ribosomal sequences using the gel plotting program pDRAW32 (http://www.acaclone.com/) and a phylogenetic tree was constructed using the neighbour-joining method of MEGA 4 (Tamura et al., 2007). BLAST analysis revealed that the Bihar phytoplasma detected in brinjal showed 98% 16S rDNA sequence identity with those of phytoplasmas from group 16SrI ('Candidatus Phytoplasma asteris'). The Bihar phytoplasma also showed only 84%, 74% and 72% 16S rDNA sequence identity respectively with those of the previously reported BLL phytoplasmas in India (EF186820, EU375486) and Bangladesh (AF228052) belonging to the 16SrVI group ('Ca. Phytoplasma trifolii'). Phylogenetic analysis (Fig. 2) evidenced that the phytoplasma associated with little leaf in brinjal in Bihar separated as a new phylogenetic branch within the 16SrI group cluster. In silico restriction fragment length polymorphism (RFLP) patterns were generated (Wei et al., 2007) for the Bihar BLL phytoplasma and the 16SrVI BLL phytoplasma reported earlier in India (EF186820) as well as the 16SrI Indian phytoplasmas identified in sandal spike (EF198362) and withania (DQ151998) (Fig. 3) with 13 restriction enzymes (AluI, BamHI, BfaI, DraI, EcoRI, HaeIII, HhaI, HinfI, HpaI, HpaII, KpnI, I, and TaqI). All the RFLP profiles of the Bihar BLL phytoplasma were similar to those of the 16SrI phytoplasmas, except for the AluI and KpnI RFLP patterns that differed from those exhibited by the 16SrVI BLL phytoplasma (EF186820). RFLP and the sequence results confirmed that the Bihar BLL phytoplasma is closely related to the phytoplasma group 16SrI and may represent a new subgroup within this group. This is the first report of a 16SrI phytoplasma affecting brinjal in India. The fact that two different phytoplasma groups (16SrVI and 16SrI) have been associated with little leaf diseases in brinjal may have further significant impact on disease epidemiology and control in India. The authors are grateful to the Department of Biotechnology, Government of India for funding.

Iron deficiency is estimated to affect over half of the world's population (Zheng et al., 2010). Wheat flour is the main ingredient of bread, and is a staple food for many civilizations. In wheat grains, iron is located in the outer layer, called bran and is lost substantially during milling and processing (Zhang et al., 2010). Thus, the flour is almost devoid of iron. Wheat grain with iron-enriched endosperm is of great interest. It is desirable to understand the bottlenecks which prevent iron translocation from the outer bran layers into the endosperm in cereals. Synchrotron radiation can be useful in identifying the transport and state of metals in plant tissues. We generated element localization maps in grains of contrasting genotypes of wheat by using synchrotron powered beam line, VESPERS (very sensitive elemental and structural probe employing radiation), at Canadian Light Source, Saskatoon, Canada. The X-ray Fluorescence analysis was focused at crease, aleurone and endosperm of mature seeds from Triticum aestivum and Aegilops kotschyi. In high iron genotypes, iron was present at higher concentration in the vascular cells than the aleurone. In contrast, in the low iron genotypes, most of the Fe was mobilized from vascular cells to aleurone cells. To further validate this result and to get accurate quantification of Fe in different tissues, micro-PIXE (Particle induced x-ray emission) analysis of the same samples is in progress at the Biotechnical Faculty and Jozef Stefan Institute, Ljubljana, Slovenia. We recorded XANES (X-ray Absorption Near Edge Structure) spectra at different points of the crease and aleurone layer. The oxidation state of iron in all cases was Fe 3+. The pre-edge structures showed one bump, suggesting tetraedric or octaedric Fe-oxygen coordination. To know the chemical coordination environment around iron centre in the cells of different tissues (such as vascular cells, transfer cells, aleurone cells, and endosperm cells), EXAFS (extended X-ray absorption fine structure) is being run on the wheat grain samples.

The relationships among diversity, productivity and climate are complex, especially in tropical ecosystems; and are less studied. We studied here the bi- and tri-partite relationships between and among the plant diversity, productivity and climate variables (i.e., temperature, precipitation and moisture) in a tropical ecosystem in India by analyzing three forest types for the year 2010. Comparison of productivity (Carnegie–Ames–Stanford Approach model derived-net primary productivity, NPP) and climate condition with respect to 2001 showed significant increase in NPP and temperature and overall decrease in precipitation and moisture condition in last decade. The NPP for sal forest ranged from 579.4 to 1,142 for the year 2001 and 557.2 to 1231.6 g Cm−2 year−1 for the year 2010. The mean monthly temperature varied from 18 to 21.8°C and 26 to 27°C; and the annual precipitation varied from 15 to 45 and 32 to 76 cm/year for the year 2001 and 2010 respectively. Strong correlation was observed between monthly mean temperature and productivity during 2001, while a decrease was observed during the year 2010. The ecosystem has shown a trend of rapid drying in last decade, triggering more detail studies for understanding of the eco-climatology. In general, sal forest, a natural forest sowed higher diversity and productivity followed by mixed deciduous (MD) forest and teak plantation. Though, areas with higher NPP are well correlated with higher temperature, precipitation and moisture; they seem to be related to species density rather than diversity. However, there is a need to have year-on-year assessments in order to ascertain if this difference represents a real phenomenon rather than an artefact of sampling. The pilot study helps in understanding the complex relationships and is very useful in the fast changing climate.

New Disease ReportsVolume 25, Issue 1 p. 9-9 ArticleOpen Access First report of Radish leaf curl virus infecting okra in India J. Kumar, Corresponding Author J. Kumar jitsingh27281@gmail.com National Agri-Food Biotechnology Institute, Mohali, 160071 Punjab, IndiaSearch for more papers by this authorA. Kumar, A. Kumar School of Biochemistry, Devi Ahilya Vishwavidyalaya, Indore, IndiaSearch for more papers by this authorS.P. Singh, S.P. Singh National Agri-Food Biotechnology Institute, Mohali, 160071 Punjab, IndiaSearch for more papers by this authorJ.K. Roy, J.K. Roy National Agri-Food Biotechnology Institute, Mohali, 160071 Punjab, IndiaSearch for more papers by this authorA. Lalit, A. Lalit Department of Biochemistry and Genetics, Barkatullah University, Bhopal, IndiaSearch for more papers by this authorD. Parmar, D. Parmar Department of Biochemistry and Genetics, Barkatullah University, Bhopal, IndiaSearch for more papers by this authorN.C. Sharma, N.C. Sharma Department of Biochemistry and Genetics, Barkatullah University, Bhopal, IndiaSearch for more papers by this authorR. Tuli, R. Tuli National Agri-Food Biotechnology Institute, Mohali, 160071 Punjab, IndiaSearch for more papers by this author J. Kumar, Corresponding Author J. Kumar jitsingh27281@gmail.com National Agri-Food Biotechnology Institute, Mohali, 160071 Punjab, IndiaSearch for more papers by this authorA. Kumar, A. Kumar School of Biochemistry, Devi Ahilya Vishwavidyalaya, Indore, IndiaSearch for more papers by this authorS.P. Singh, S.P. Singh National Agri-Food Biotechnology Institute, Mohali, 160071 Punjab, IndiaSearch for more papers by this authorJ.K. Roy, J.K. Roy National Agri-Food Biotechnology Institute, Mohali, 160071 Punjab, IndiaSearch for more papers by this authorA. Lalit, A. Lalit Department of Biochemistry and Genetics, Barkatullah University, Bhopal, IndiaSearch for more papers by this authorD. Parmar, D. Parmar Department of Biochemistry and Genetics, Barkatullah University, Bhopal, IndiaSearch for more papers by this authorN.C. Sharma, N.C. Sharma Department of Biochemistry and Genetics, Barkatullah University, Bhopal, IndiaSearch for more papers by this authorR. Tuli, R. Tuli National Agri-Food Biotechnology Institute, Mohali, 160071 Punjab, IndiaSearch for more papers by this author First published: 01 March 2012 https://doi.org/10.5197/j.2044-0588.2012.025.009Citations: 9AboutSectionsPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onFacebookTwitterLinkedInRedditWechat Okra (Abelmoschus esculentus) is one of the important vegetable crops of India cultivated in 452 hectares and yielding 4803 tonnes (Anonymous, 2010). In Bihar State, India, leaf curl disease on okra was observed in the field causing crop losses of about 30% in 2009 and 35% in 2010-11. Characteristic symptoms of this disease were leaf curling and overall stunting of plants that bore no fruit (Fig. 1A). To test for a begomovirus-infection, total DNA was extracted from symptom-bearing leaves of six infected plants (2 plants ' 3 fields). A PCR approach was used to amplify viral genomes (primers F1For/Rev and F2For/Rev; Kumar et al., 2011) or alpha- and betasatellites (primers 'nanofor'/'nanorev' and 01/04; Kumar et al., 2010). All samples yielded PCR-products for a begomovirus and associated satellites, which were cloned and sequenced. The sequence deposited in GenBank for the monopartite begomovirus (Accession No. HQ257375) showed 97% and 92% nucleotide identity to Radish leaf curl virus (RaLCV) depositions GU732203 and EF175733, respectively. The alphasatellite (Accession No. HQ728354) possessed 98% and 96% nucleotide identity to Cotton leaf curl Burewala alphasatellite (CLCuBwA) sequences HM004548 and FN658728, respectively. The betasatellite (Accession No. HQ257376) exhibited 96% and 94% nucleotide identity to Tomato leaf curl Bangladesh betasatellite (ToLCBDB) sequences GU732208 and EF190215, respectively. Phylogenetic analysis of the begomovirus genome revealed a close relationship with RaLCV,GU732203 and EF175733 but a distant relationship with other okra infecting begomoviruses in India, AF241479 and FJ176236 (Fig. 2A). Phylogenetic analysis of the alphasatellite showed close relationship with CLCuBwA (Fig. 2B) and the betasatellite with ToLCBDB (Fig. 2C). Figure 1Open in figure viewerPowerPoint Figure 2Open in figure viewerPowerPoint Rolling circle amplification (RCA) was performed (TempliPhi amplification kit; GE Healthcare, USA) to construct infectious clones. RCA products were partially digested with BamHI to obtain monomer and head-to-tail tandem repeat dimers of full-length begomoviral DNA. Monomers and dimers were cloned into the pCAMBIA1301 vector. Infectious head-to-tail tandem repeat clones of both satellites were prepared analogously. Sequencing of 30 begomoviral monomer clones confirmed the presence of identical viral DNAs. Healthy whiteflies (~25) were used for virus and satellite transmission from field-collected infected plants to healthy tobacco and okra. Infectivity testing was performed by inoculation of tobacco and okra (10 plants each) with a mixture of begomovirus and alpha- and betasatellite infectious clones. Plants of both assays yielded typical symptoms of leaf curling and stunting identical to those observed previously in the field. Thus RaLCV and its associated satellites were confirmed as the causal agent of okra leaf curl disease. However, the functional role of both satellites for symptom development remains to be determined. This is the first report providing the evidence for RaLCVinfecting okra in India. Acknowledgments Authors are grateful to the Executive Director, NABI for providing facilities. Thanks to Dr. A K Pandey for critical reading of the manuscript. References Anonymous, 2010. Okra production status by Indian Horticulture database, National Horticulture Board. [http://nhb.gov.in/area-pro/ch2.pdf]. Kumar J, Kumar A, Roy JK, Tuli R, Khan JA, 2010. Identification and molecular characterization of begomovirus and associated satellite DNA molecules infecting Cyamopsis tetragonoloba. Virus Genes 41, 118– 125. [doi:10.1007/s11262-010-0482-7] Kumar A, Snehi SK, Raj SK, Kumar J, Khan JA, 2011. Association of Cotton leaf curl Burewala virus and its satellite molecules with leaf distortion symptoms of cotton in India. New Disease Reports 24, 18. [doi:10.5197/j.2044-0588.2011.024.018] Citing Literature Volume25, Issue1January 2012-June 2012Pages 9-9 FiguresReferencesRelatedInformation

Monopartite begomoviruses comprise DNA-A as the main genome and associated satellite DNAs. Viral DNA extracted from guar (Cyamopsis tetragonoloba) showing leaf curl symptoms exhibited positive amplification of coat protein (CP) gene of DNA-A component, suggesting the presence of begomovirus. Full length DNA-A was amplified by primer pair re-designed from CP gene nucleotide sequence. The associated alphasatellite and betasatellite DNA molecules were amplified and sequenced, confirming the presence of monopartite begomovirus. Sequence comparisons showed 89% identity with other begomoviruses. The Neighbor-Joining tree based on full length DNA-A nucleotide sequence showed that the guar infecting begomovirus clustered separately from other known begomoviruses. The betasatellite shared a high (96%) nucleotide identity to Cotton leaf curl Multan betasatellites. The alphasatellite showed 91% nucleotide identity to alphasatellite associated with begomovirus infecting Okra. Recombination analyses showed three recombinant fragments in DNA-A, two in betasatellite, and four in alphasatellite. The results suggest that the begomovirus identified in this study was a new recombinant virus. Its name was proposed as Cyamopsis tetragonoloba leaf curl virus (CyTLCuV).

Simulated pot experiments were conducted on four rice (Oryza sativa L.) genotypes (Triguna, IR-36, PNR-519, and IET-4786) to examine the effects of As(V) on amino acids and mineral nutrient status in grain along with antioxidant response to arsenic exposure. Rice genotypes responded differentially to As(V) exposure in terms of amino acids and antioxidant profiles. Total amino acid content in grains of all rice genotypes was positively correlated with arsenic accumulation. While, most of the essential amino acids increased in all cultivars except IR-36, glutamic acid and glycine increased in IET-4786 and PNR-519. The level of nonprotein thiols (NPTs) and the activities of superoxide dismutase (SOD; EC 1.15.1.1), glutathione reductase (GR; EC 1.6.4.2) and ascorbate peroxidase (APX; EC 1.11.1.11) increased in all rice cultivars except IET-4786. A significant genotypic variation was also observed in specific arsenic uptake (SAU; mg kg(-1)dw), which was in the order of Triguna (134) > IR-36 (71) > PNR-519 (53) > IET-4786 (29). Further, application of As(V) at lower doses (4 and 8 mg L(-1) As) enhanced the accumulation of selenium (Se) and other nutrients (Fe, P, Zn, and S), however, higher dose (12 mg L(-1) As) limits the nutrient uptake in rice. In conclusion, low As accumulating genotype, IET-4786, which also had significantly induced level of essential amino acids, seems suitable for cultivation in moderately As contaminated soil and would be safe for human consumption.

Very little is known about molecular events associated with callus differentiation in indica rice. The genes expressed differentially during shoot meristem initiation were identified on genomic arrays applied to efficiently regenerating rice calli. A thidiazuron (TDZ; N-phenyl-N-thiadiazol-1,2,3-5,ylurea)-dependent regeneration protocol was developed for efficient embryogenesis in indica rice. The regenerating embryogenic calli induced by TDZ for 10 days showed transcriptional modulation of a number of genes associated with photosynthesis, hormone metabolism, plant development, signal transduction, light response, and plant defense. Eighteen candidate miRNAs were predicted to target the genes expressed differentially in the embryogenic calli grown in TDZ-containing medium. The majority of the photosynthesis-related genes up-regulated in differentiating calli were not expressed or were down-regulated in developing seeds and inflorescences. Most of the genes down-regulated in differentiating calli were up-regulated in developing seeds. The transcriptome of differentiating callus most closely resembled that of the germinating whole seed.

The exposure of paddy fields to arsenic (As) through groundwater irrigation is a serious concern that may not only lead to As accumulation to unacceptable levels but also interfere with mineral nutrients in rice grains. In the present field study, profiling of the mineral nutrients (iron (Fe), phosphorous, zinc, and selenium (Se)) was done in various rice genotypes with respect to As accumulation. A significant genotypic variation was observed in elemental retention on root Fe plaque and their accumulation in various plant parts including grains, specific As uptake (29-167 mg kg(-1) dw), as well as As transfer factor (4-45%). Grains retained the least level of As (0.7-3%) with inorganic As species being the dominant forms, while organic As species, viz., dimethylarsinic acid and monomethylarsonic acid, were non-detectable. In all tested varieties, the level of Se was low (0.05-0.12 mg kg(-1) dw), whereas that of As was high (0.4-1.68 mg kg(-1) dw), considering their safe/recommended daily intake limits, which may not warrant their human consumption. Hence, their utilization may increase the risk of arsenicosis, when grown in As-contaminated areas.

ABSTRACT An efficient method of in vitro shoot propagation of six elite accessions of Withania somnifera collected throughout India was developed. Maximum numbers of shoots in all accessions were achieved from axillary explant on Murashige and Skoog (MS) medium supplemented with 1 mgL−1 BAP and 1 mgL−1 kinetin. Inclusion of kinetin increased shoot numbers in a shorter time-period and was effective on all the elite accessions. The highest number of shoots (60 ± 1.82 and 60.05 ± 2.03) was observed in two lines developed by CIMAP, Lucknow, while the lowest number of shoots (19.4 ± 1.63) occurred on a wild collected accession with bushy growth. The in vitro raised shoots of all the accessions could be easily rooted on MS medium supplemented with 2 mgL−1 IBA. Rooted shoots were successfully established in a garden soil-vermi-compost (3:1, w:w) medium in a glasshouse. Stable production of withanolides from in vitro regenerated shoots was comparable to the yields from field grown mother plants, indicating the in vitro methodology could be used successfully for the true-to-type plant regeneration of Withania somnifera accessions.

olated promising genes and promoters, with potential for commercialization. However, isolated growth of researchers, limited R&D infrastructure and knowledge base available in the seed industr y, and the regulatory processes involved in the release of transgenic cultivars pose serious challenges to accelerating the conversion of such leads into field crops. There is an urgent need to evolve efficient managerial approaches for developing and advancing genetically engineered cultivars into Indian agriculture. This article emphasizes the need to identify the most promising transgenics and genes available in the country, evaluate the related intellectual property issues and provide unstinted support to accelerate the process of their commercialization. An approach that would sy nergize public–public and public–private partnerships needs to be evolved. Initially, it may be driven by public sector through crop- and trait-specific consortia, comprising researchers and the seed industry. A few of such examples should be taken to the stage of clearance by GEAC for unreg ulated release of indigenously developed transgenic cu ltivars. These would then serve as role models and catalyse the formation of need -based teams and lasting partnerships, needed to usher Indian agriculture into a globally competitive phase of su stainable productivity. IT has been amply demonstrated that genetically engineered (GE) seeds, developed using recombinant-DNA techniques (also referred to as transgenics, or genetically modified organisms (GMOs)), provide an easily adaptable, scaleneutral technology for the farmers. Even farmers with small land holdings can benefit from the high technology when such seeds become available to them at reasonable cost. Changes required in the existing crop management practices, or additional inputs are minimal. The Task Force on Agri cultural Biotechnology constituted recently by the Ministry of Agriculture recommended 1

Efficient transformation of embryonal segments of pigeon pea (Cajanus cajan (L.) Millsp.) was obtained by using Agrobacterium tumefaciens strain GV2260 harboring a modified binary vector pPK202 carrying the marker gene neomycin phosphotransferase II (npt II) and a synthetic cry I E-C gene under a constitutive 35S promoter. Shoots developed on Agrobacterium treated explants were selected on MS medium supplemented with 2.0mgl−1 BAP, 250mgl−1 cefotaxime and 75mgl−1 kanamycin. Elongated kanamycin resistant shoots were subsequently rooted on MS medium supplemented with 1.0mgl−1 NAA and later transferred to sterile vermiculite followed by transfer to the transgenic green house. Integration of T-DNA into nuclear genome of transformed plants and its sexual transmission to the progeny of the transgenic plants were confirmed by PCR amplification of 700bp npt II fragment and Southern blot hybridization analysis using the PCR amplified npt II fragment as probe. In vitro insect bioassay using Spodoptera litura larvae of first and second instar stages on T1 and T2 plants shows that the expression of the synthetic cry I E-C in transgenic pigeon pea plants confers protection against the insect larvae. Western analysis showed a 71.5kDa band confirming the presence of cry I E-C protein in the T1 and T2 transgenic plants. This protocol allows effective transformation and quick regeneration of insect resistant transgenic plants of pigeon pea.

A synthetic gene coding for the surface glycoprotein (G protein) of rabies virus was strategically designed to achieve high-level expression in transgenic plants. The native signal peptide was replaced by that of the pathogenesis related protein, PR-S of Nicotiana tabacum. An endoplasmic reticulum retention signal was included at C-terminus of the G protein. Tobacco plants were genetically engineered by nuclear transformation. Selected transgenic lines expressed the chimeric G protein at 0.38% of the total soluble leaf protein. Mice immunized intraperitoneally with the G protein purified from tobacco leaf microsomal fraction elicited high level of immune response as compared to the inactivated commercial viral vaccine. The plant-derived G protein induced complete protective immunity in mice against intracerebral lethal challenge with live rabies virus. The results establish that plants can provide a safe and effective production system for the expression of immunoprotective rabies virus surface protein.

Withania somnifera (Hindi - Ashwagandha, English - winter cherry) is used in Ayurvedic formulations for a variety of health-promoting effects. Several mono- and poly-herbal products commercially available in the Indian market were quantitatively analysed for a number of chemical constituents. The results. revealed wide variations in the content of all seven constituents tested. More than 70-fold variation in the daily intake of withaferin A (the main active constituent of Ashwagandha) was found in the products. The study thus emphasizes the need for stringent phytochemical standardization of herbal products.

Nucleotide positions conserved on the 3' side of the initiator codon ATG and the corresponding N-terminal amino acid residues in a number of highly abundant plant proteins were identified by computational analysis of a dataset of highly expressed plant genes. The reporter genes uidA and gfp were modified to introduce these features. Insertion of GCT TCC TCC after the initiator codon ATG augmented expression for both the reporter genes. The insertion of each successive codon improved the expression of beta-glucuronidase (GUS) in an incremental fashion in transient transformation of tobacco (Nicotiana tabacum) leaves. The insertion of alanine-serine (Ser)-Ser resulted in about a 2-fold increase in the stability of GUS. However, this did not account for the 30- to 40-fold increase in GUS activity between the constructs coding for methionine-alanine-Ser-Ser-GUS and the native enzyme. Substitution of the codon for Ser at the third amino acid residue with synonymous codons reduced GUS expression. The results suggest a role for the conserved nucleotides in the +4 to +11 region in augmenting posttranscriptional events in the expression of genes in plants.

A dataset of highly expressed plant genes was developed from the nucleic acids sequence database. The characteristic features of the nucleotide sequences in TATA-box, transcription initiation, untranslated leader and translation initiation regions in the highly expressible genes in plants and the conserved sequences present 500 bp upstream of transcription initiation site were identified. These features were employed to theoretically design a ’minimal expression cassette’ and a promoter-upstream ’activation module.’ The ’minimal expression cassette’ was sufficient to express the gusA reporter gene in transient transformation of tobacco leaf. The context on the 3′ side of the initiator codon, conserved in a majority of the highly expressible genes, gave approximately a ninefold increase in the expression of β-glucuro- nidase. The artificially designed, upstream ’activation module’ enhanced gusA expression further by about 30-fold in transiently transformed tobacco leaves. A 450-bp-long complete expression cassette, containing both the ’minimal expression cassette’and the ’activation module’ expressed gusA at a high level in cotton leaves, potato tubers and cabbage stem also. In stably transformed tobacco plants, the ’complete expression cassette’ expressed gusA at levels higher than the native CaMV 35S promoter. Histological studies established that the ’complete expression cassette’ was expressed at a high level in different cell types in the roots, leaves, vascular tissues and flower parts of the transgenic tobacco plants. The results substantiate the functional validity of the features identified by us and demonstrate the potential of computational biology in designing artificial expression cassettes for applications in biotechnology.

A direct and indirect methodology for efficient shoot proliferation and regeneration from various explants of Mentha arvensis has been developed by studying the interactive effect of plant growth regulator and light colour. Nodal explants cultured on Murashige and Skoog's (MS) medium supplemented with 10 muM thidiazuron and incubated under red light, proliferated an average of 50 shoots after 30 days, whereas internodal explants cultured in modified MS medium supplemented with 40 muM 6-benzylaminopurine and 0.5 muM alpha -naphthaleneacetic acid and exposed to red light, regenerated an average of 200 shoots after 60 days. Among the six cultivars tested, internodal explants of cv, Gomti regenerated about 90 shoots after 20 days of culture. Under the above conditions, leaf explants were observed to regenerate an average of 60 shoots after 60 days of culture, which was preceded by callus development. The protocol standardized for high efficiency shoot proliferation and regeneration in M, arvensis from nodal, internodal and leaf explants is suitable for micropropagation, genetic transformation and for obtaining somaclonal variants in this essential oil-yielding crop plant.

The filamentous non-heterocystous cyanobacterium Plectonema boryanum fixes dinitrogen at a high rate during microaerobic growth in continuous illumination by temporal separation of oxygen-evolving photosynthesis and oxygen-sensitive dinitrogen fixation. The onset of nitrogen fixation is preceded by a depression in photosynthesis that establishes a sufficiently low level of dissolved oxygen in the growth medium. A several-fold reduction in the level of transcripts coding for phycocyanin (cpcBA) and the chlorophyll a binding protein of photosystem II (psbC) and psbA accompanied the depression in photosynthetic oxygen evolution. Unlike most of the other organisms examined to date, in P. boryanum, psbC and psbD do not appear to be co-transcribed. The psbC transcripts were down-regulated several fold, while the psbD transcript declined marginally during the nitrogen fixation phase. A decrease in dissolved oxygen and a dramatic increase in the level of nifH transcripts and the enzyme activity of nitrogenase were characteristic of the nitrogen fixation phase. The level of transcript for glnA, which encodes glutamine synthetase, was not altered. Reciprocal regulation of gene expression was well orchestrated with the alternating cycles of photosynthesis and nitrogen fixation in P. boryanum.

Cotyledonary nodes taken alongwith shoot apex from seedlings of cotton (G. hirsutum) proliferated into shoots on nutrient agar medium supplemented with cytokinins. In the presence of optimal plant growth regulators, low light intensity enhanced the number of shoots initiated per explant in cotton. An average of 33.5 +/- 2.9 shoots were obtained from a single explant cultured for 8 weeks which is about four fold higher than the values reported in earlier protocols. The isolated shoots were rooted on nutrient agar medium supplemented with alpha-naphthalene acetic acid and transferred to soil after acclimatization. Regenerated plants were morphologically identical to the seed-germinated plants and were fertile.