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Computational analysis of common bean (Phaseolus vulgaris L., genotype BAT93) lycopene ?-cyclase and ?-carotene hydroxylase gene's cDNA.

ABSTRACT: The identification of genes and understanding of genes' expression and regulation in common bean (Phaseolus vulgaris L.) is necessary in order to strategize its improvement using genetic engineering techniques. Generation of expressed sequence tags (ESTs) is useful in rapid isolation, identification and characterization of the genes. To study the gene expression in P. vulgaris pods tissue, ESTs generation work was initiated. Early stage and late stage bean-pod-tissues cDNA libraries were constructed using CloneMiner cDNA library construction kit. In total, 5972 EST clones were isolated using random method of gene isolation. While processing ESTs, we found lycopene ?-cyclase (PvLCY-?) and ?-carotene hydroxylase (PvCHY-?) gene's cDNA. In carotenoid biosynthesis pathway, PvLCY-? catalyzes the production of carotene; and PvCHY-? is known to function as a catalyst in the production of lutein and zeaxanthin. To understand more about PvLCY-? and PvCHY-?, both strands of both cDNA clones were sequenced using M13 forward and reverse primers. Nucleotide and deduced protein sequences were analyzed and annotated using online bioinformatics tools. Results showed that PvLCY-? and PvCHY-? cDNAs are 1639 and 1107 bp in length, respectively. Analysis results showed that PvLCY-? and PvCHY-? gene's cDNA contains an open reading frame (ORF) that encodes for 502 and 305 amino acid residues, respectively. The deduced protein sequence analysis results also showed the presence of conserved domains needed for PvLCY-? and PvCHY-? functions. The phylogenetic analysis of both PvLCY-? and PvCHY-? proteins showed it's closeness with the LCY-? and CHY-? proteins from Glycine max, respectively. The nucleotide sequence of PvLCY-? and PvCHY-? gene's cDNA and it's annotation is reported in this paper.

SUBMITTER: Bhore SJ

PROVIDER: S-EPMC3602890 | biostudies-literature | 2013

REPOSITORIES: biostudies-literature

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Computational analysis of common bean (Phaseolus vulgaris L., genotype BAT93) lycopene β-cyclase and β-carotene hydroxylase gene's cDNA.

Bhore Subhash Janardhan SJ Amelia Kassim K Wang Edina E Priyadharsini Sindhuja S Shah Farida Habib FH

Bioinformation 20130221 4

The identification of genes and understanding of genes' expression and regulation in common bean (Phaseolus vulgaris L.) is necessary in order to strategize its improvement using genetic engineering techniques. Generation of expressed sequence tags (ESTs) is useful in rapid isolation, identification and characterization of the genes. To study the gene expression in P. vulgaris pods tissue, ESTs generation work was initiated. Early stage and late stage bean-pod-tissues cDNA libraries were constru ...[more]

PMID: 23519320

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Project description:Traditional yellow maize though contains high kernel carotenoids, the concentration of provitamin A (proA) is quite low (<2 μg/g), compared to recommended level (15 μg/g). It also possesses poor endosperm protein quality due to low concentration of lysine and tryptophan. Natural variant of crtRB1 (β-carotene hydroxylase) and lcyE (lycopene-ε-cyclase) cause significant enhancement of proA concentration, while recessive allele, opaque2 (o2) enhances the level of these amino acids. Development of biofortified maize enriched in proA, lysine and tryptophan thus holds significance in alleviation of micronutrient malnutrition. In the present study, marker-assisted stacking of crtRB1, lcyE and o2 was undertaken in the genetic background of four maize hybrids (HQPM1, HQPM4, HQPM5, and HQPM7) popularly grown in India. HP704-22 and HP704-23 were used as donors, while four elite QPM parents viz., HKI161, HKI163, HKI193-1, and HKI193-2 were used as recipients. CrtRB1 showed severe segregation distortion, while lcyE segregated as per the expectation. Recovery of recurrent parent genome (RPG) among selected backcross progenies ranged from 89 to 93%. Introgressed progenies possessed high concentration of proA (7.38-13.59 μg/g), compared to 1.65-2.04 μg/g in the recurrent parents. The reconstituted hybrids showed an average of 4.5-fold increase in proA with a range of 9.25-12.88 μg/g, compared to original hybrids (2.14-2.48 μg/g). Similar plant-, ear-, and grain- characteristics of improved versions of both inbreds and hybrids were observed when evaluated with their respective original versions. Mean lysine (0.334%) and tryptophan (0.080%) of the improved hybrids were at par with the original versions (lysine: 0.340%, tryptophan: 0.083%). Improved hybrids also possessed similar grain yield potential (6,301-8,545 kg/ha) with their original versions (6,135-8,479 kg/ha) evaluated at two locations. This is the first study of staking crtRB1-, lcyE-, and o2-, favorable alleles in single genetic background. The improved inbreds can be effectively used as potential donor for independent and/or simultaneous introgression of crtRB1, lcyE, and o2 in the future breeding programme. These biofortified maize hybrids, rich in proA, lysine and tryptophan will hold great promise for nutritional security.

Project description:Common bean (Phaseolus vulgaris L.) is a staple food in Brazil with both nutritional and socioeconomic importance. As an orphan crop, it has not received as much research attention as the commodity crops. Crop losses are strongly related to virus diseases transmitted by the whitefly Bemisia tabaci, one of the most important agricultural pests in the world. The main method of managing whitefly-transmitted viruses has been the application of insecticides to reduce vector populations. Compared to chemical vector control, a more sustainable strategy for managing insect-borne viruses is the development of resistant/tolerant cultivars. RNA interference has been applied to develop plant lines resistant to the whitefly in other species, such as tomato, lettuce and tobacco. Still, no whitefly-resistant plant has been made commercially available to date. Common bean is a recalcitrant species to in vitro regeneration; therefore, stable genetic transformation of this plant has been achieved only at low frequencies (<1%) using particle bombardment. In the present work, two transgenic common bean lines were obtained with an intron-hairpin construct to induce post-transcriptional gene silencing against the B. tabaci vATPase (Bt-vATPase) gene, with stable expression of siRNA. Northern blot analysis revealed the presence of bands of expected size for siRNA in leaf samples of the line Bt-22.5, while in the other line (11.5), the amount of siRNA produced was significantly smaller. Bioassays were conducted with both lines, but only the line Bt-22.5 was associated with significant mortality of adult insects (97% when insects were fed on detached leaves and 59% on the whole plant). The expression of the Bt-vATPase gene was 50% lower (p < 0.05) in insects that fed on the transgenic line Bt-22.5, when compared to non-transgenic controls. The transgenic line did not affect the virus transmission ability of the insects. Moreover, no effect was observed on the reproduction of non-target organisms, such as the black aphid Aphis craccivora, the leafminer Liriomyza sp. and the whitefly parasitoid Encarsia formosa. The results presented here serve as a basis for the development of whitefly-tolerant transgenic elite common bean cultivars, with potential to contribute to the management of the whitefly and virus diseases.

Dataset Information

Computational analysis of common bean (Phaseolus vulgaris L., genotype BAT93) lycopene ?-cyclase and ?-carotene hydroxylase gene's cDNA.

Publications

Computational analysis of common bean (Phaseolus vulgaris L., genotype BAT93) lycopene β-cyclase and β-carotene hydroxylase gene's cDNA.

Similar Datasets

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