@article { ISI:000259484700016, title = {Direct shoot organogenesis and plant regeneration from hypocotyl explants in selected genotypes of Leucaena leucocephala - a leguminous pulpwood tree}, journal = {Indian Journal of Biotechnology}, volume = {7}, number = {3}, year = {2008}, month = {JUL}, pages = {388-393}, publisher = {NATL INST SCIENCE COMMUNICATION-NISCAIR}, type = {Article}, address = {DR K S KRISHNAN MARG, PUSA CAMPUS, NEW DELHI 110 012, INDIA}, abstract = {

An efficient in vitro plant regeneration system in subabul (Leucaena leucocephala), a leguminous pulp wood tree species, was established. The induction of shoots was achieved from selected elite clones of subabul K-8, K-636 and also wild type on MS medium supplemented with 2\% sucrose and different concentrations (0.88 to 24.6 mu M) of plant growth regulators (BA, Kn, 2iP \& TDZ). The best medium for shoot regeneration was MS with 22.2 mu M BA (5 shoots per explant), followed by 22.7 mu M TDZ (4.6 shoots per explant). Addition of putriscine (9.3 mu M) to MS medium containing 22.2 mu M BA enhanced the number of multiple shoots to 7-8 but not the frequency of response. Shoot initials (measuring 1 cm) when separated and transferred on to MS medium containing 1.4 mu M GA(3) elongated to 2-5 cm in 15.20 d with 80\% frequency. The per cent frequency of shoot differentiation was almost identical in the genotypes K-8 and K-636 but it differed significantly from the wild type. Leaf yellowing and abscission in all the genotypes was curtailed by supplementing the medium with 685 mu M glutamine or 540 mu M adenine. The excised shoots were transferred to root regeneration media containing 2.46 and 4.98 mu M IBA or 2.6 and 5.3 mu M NAA. Root regeneration was noticed with 100\% frequency in all the three genotypes in presence of IBA or NAA. Plantlets were transferred successfully to the pots with 70\% survival rate with no visible morphological variations. The protocol can be utilized for mass propagation and genetic transformation studies of this important pulpwood species.

}, keywords = {hypocotyl explants, Leucaena lucocephala, pulpwood leguminous tree, shoot organogenesis}, issn = {0972-5849}, author = {Sirisha, V. L. and Prashant, S. and Ranadheer, D. and Ramprasad, P. and Shaik, N. M. and Arha, Manish and Gupta, S. K. and Srivastava, Sarneer and Yadav, A. K. and Kulkarni, P. S. and Othalathara U. Abhilash and Khan, Bashir Mohammad and Rawal, Shuban K. and Kishor, P. B. Kavi} } @article { ISI:000257961500018, title = {Isolation and heterologous expression of PHA synthesising genes from Bacillus thuringiensis R1}, journal = {World Journal of Microbiology \& Biotechnology}, volume = {24}, number = {9}, year = {2008}, month = {SEP}, pages = {1769-1774}, publisher = {SPRINGER}, type = {Article}, address = {233 SPRING ST, NEW YORK, NY 10013 USA}, abstract = {

The polyhydroxyalkanoate biosynthesis gene locus from Bacillus thuringiensis R1 was isolated, cloned and analyzed at the molecular level. We found that a similar to 5 kb SacI-ClaI digested fragment of genomic DNA from B. thuringiensis R1 encoding the PHA synthesising genes, conferred PHA producing ability to E. coli. The fragment was sequenced and found to be of 4787 bp with five open reading frames. Sequence alignment with closely related species of Bacillus in the existing database revealed that the ORFs correspond to phaP, phaQ, phaR, phaB and phaC genes. However, E. coli harboring phaP, phaQ, phaR, phaB and phaC locus produced very low PHA. Furthermore, complementation of the locus with phaA from Ralstonia eutropha increased the PHA production in the recombinant E. coli from 3.0\% to 24\% of cell dry mass. The putative promoter regions and ribosome binding sites were identified for each of the gene. Conserved domains for PHA synthase and aceto-acetyl-coA reductase were also identified. We hence conclude that the PHA operon of Bacillus thuringiensis R1 consists of phaP, phaQ, phaR, phaB, phaC and complementation of the same with phaA is accountable for its high PHA production.

}, keywords = {aceto-acetyl-coA reductase, Bacillus thuringiensis R1, PHA synthase, polyhydroxyalkanoate}, issn = {0959-3993}, doi = {10.1007/s11274-008-9669-7}, author = {Desetty, Rohini D. and Mahajan, Vineet S. and Khan, Bashir Mohammad and Rawal, Shuban K.} } @article { ISI:000287716400004, title = {Expression analysis of cinnamoyl-CoA reductase (CCR) gene in developing seedlings of Leucaena leucocephala: A pulp yielding tree species}, journal = {Plant Physiology and Biochemistry}, volume = {49}, number = {2}, year = {2011}, month = {FEB}, pages = {138-145}, publisher = {ELSEVIER FRANCE-EDITIONS SCIENTIFIQUES MEDICALES ELSEVIER}, address = {23 RUE LINOIS, 75724 PARIS, FRANCE}, abstract = {

Removal of lignin is a major hurdle for obtaining good quality pulp. Leucaena leucocephala (subabul) is extensively used in paper industry in India; therefore, as a first step to generate transgenic plants with low lignin content, cDNA and genomic clones of CCR gene were isolated and characterized. The cDNA encoding CCR (EC 1.2.1.44) was designated as LI-CCR; the sequence analysis revealed an Open Reading Frame (ORF) of 1005 bp. Phylogenetic analysis showed that LI-CCR sequence is highly homologous to CCRs from other dicot plants. The 2992 bp genomic clone of Leucaena CCR consists of 5 exons and 4 introns. The haploid genome of L leucocephala contains two copies as revealed by DNA blot hybridization. LI-CCR gene was over-expressed in Escherichia coli, which showed a molecular mass of approximately 38 kDa. Protein blot analysis revealed that LI-CCR protein is expressed at higher levels in root and in stem, but undetectable in leaf tissues. Expression of CCR gene in Leucaena increased up to 15 d in case of roots and stem as revealed by QRT-PcR studies in 0-15 d old seedlings. ELISA based studies of extractable CCR protein corroborated with QRT-PCR data. CCR protein was immuno-cytolocalized around xylem tissue. Lignin estimation and expression studies of 5,10 and 15 d old stem and root suggest that CCR expression correlates with quantity of lignin produced, which makes it a good target for antisense down regulation for producing designer species for paper industry. (C) 2010 Elsevier Masson SAS. All rights reserved.

}, keywords = {Cinnamoyl-CoA reductase, Developing seedling, Leucaena leucocephala, Lignin biosynthesis}, issn = {0981-9428}, doi = {10.1016/j.plaphy.2010.11.001}, author = {Srivastava, Sameer and Gupta, Ranadheer K. and Arha, Manish and Vishwakarma, Rishi K. and Rawal, Shuban K. and Kishor, P. B. Kavi and Khan, Bashir Mohammad} } @article { ISI:000313165500067, title = {Functional characterization, homology modeling and docking studies of beta-glucosidase responsible for bioactivation of cyanogenic hydroxynitrile glucosides from Leucaena leucocephala (subabul)}, journal = {Molecular Biology Reports}, volume = {40}, number = {2}, year = {2013}, month = {FEB}, pages = {1351-1363}, publisher = {SPRINGER}, address = {VAN GODEWIJCKSTRAAT 30, 3311 GZ DORDRECHT, NETHERLANDS}, abstract = {

Glycosyl hydrolase family 1 beta-glucosidases are important enzymes that serve many diverse functions in plants including defense, whereby hydrolyzing the defensive compounds such as hydroxynitrile glucosides. A hydroxynitrile glucoside cleaving beta-glucosidase gene (Llbglu1) was isolated from Leucaena leucocephala, cloned into pET-28a (+) and expressed in E. coli BL21 (DE3) cells. The recombinant enzyme was purified by Ni-NTA affinity chromatography. The optimal temperature and pH for this beta-glucosidase were found to be 45 A degrees C and 4.8, respectively. The purified Llbglu1 enzyme hydrolyzed the synthetic glycosides, pNPGlucoside (pNPGlc) and pNPGalactoside (pNPGal). Also, the enzyme hydrolyzed amygdalin, a hydroxynitrile glycoside and a few of the tested flavonoid and isoflavonoid glucosides. The kinetic parameters K (m) and V (max) were found to be 38.59 mu M and 0.8237 mu M/mg/min for pNPGlc, whereas for pNPGal the values were observed as 1845 mu M and 0.1037 mu M/mg/min. In the present study, a three dimensional (3D) model of the Llbglu1 was built by MODELLER software to find out the substrate binding sites and the quality of the model was examined using the program PROCHEK. Docking studies indicated that conserved active site residues are Glu 199, Glu 413, His 153, Asn 198, Val 270, Asn 340, and Trp 462. Docking of rhodiocyanoside A with the modeled Llbglu1 resulted in a binding with free energy change (Delta G) of -5.52 kcal/mol on which basis rhodiocyanoside A could be considered as a potential substrate.

}, keywords = {Glycosyl hydrolase family 1, homology modeling, Leucaena leucocephala, Molecular docking}, issn = {0301-4851}, doi = {10.1007/s11033-012-2179-6}, author = {Shaik, Noor M. and Misra, Anurag and Singh, Somesh and Fatangare, Amol B. and Ramakumar, Suryanarayanarao and Rawal, Shuban K. and Khan, Bashir Mohammad} }