Indian Journal of Biotechnology


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ISSN:0972-5849                                                                            CODEN:IJBNAR 4(3) 301-428

                                                                                                                                                2005 

VOLUME 4                                                NUMBER 3                                                 JULY 2005

 

 

CONTENTS

 

Reviews

 

 

FISH and GISH: Modern cytogenetic techniques

307

 

        IPC Code: Int. Cl.7 A01H1/00

 

 

        J Devi, J M Ko & B B Seo

 

 

 

 

 

RNA Interference: A revolution in drug development

316

 

        IPC Code: Int. Cl.7 A61K31/713, 48/00

 

 

        Neha Gairola, Sonali N Joshi & Suneela S Dhaneshwar

 

 

 

 

 

Papers

 

 

 

 

 

Analysis of PCR products for PHB production in indigenous Pseudomonas sp. LDC-5

323

 

 IPC Code: Int. Cl.7 C08G63/00; C12N15/10; C12R1:38

 

 

        K Sujatha, R Shenbagarathai & A Mahalakshmi

 

 

 

 

 

Partial purification and characterization of a-amylase produced by Aspergillus oryzae using spent-brewing grains

 

336

 

        IPC Code: Int. Cl.7 A01N63/02; C12N9/30; C12R1:69

 

 

Anil Kumar Patel, K Madhavan Nampoothiri, Sumitra Ramachandran, George Szakacs & Ashok Pandey

 

 

 

 

 

Screening and interaction effects of physical parameters, total N content and buffer on L(+) Lactic acid production in SSF by Lactobacillus amylophilus GV6 using Taguchi designs

 

 

342

 

        IPC Code: Int. Cl.7 A01N63/02; C12N9/00; C12R1:225

 

 

        B J Naveena, Md Altaf, K Bhadrayya & Gopal Reddy

 

 

 

 

 

Cyclodextrin glycosyl transferases from Bacillus circulance and Bacillus sp.

347

 

        IPC Code: Int. Cl.7 A01N63/02; C12N9/24; C12R1:07, 1:09

 

 

        R S Prakasham, R Sreenivas Rao, Ch Subba Rao & P N Sarma

 

 

 

 

 

Influence of growth factors on carotenoid pigmentation of Rhodotorula glutinis
DFR-PDY from natural source

 

353

 

        IPC Code: Int. Cl.7 A01N63/02; B01D:15/08; C12P23/00; C12R1:645

 

 

        B V Latha, K Jeevaratnam, H S Murali & K S Manja

 

 

.

 

 

Analysis of VNTR loci, APOB 3˘ HVR and D1S80 in North Indians

358

 

        IPC Code: Int. Cl.7 C12N15/10

 

 

        Monisha Mukherjee, Anvesha Srivastava, Akanchha Kesari & Balraj Mittal

 

 

Cloning, characterization and expression of bovine (Bos indicus) tumour necrosis factor-a

363

 

        IPC Code: Int. Cl.7 C12N15/10, 15/28

 

 

        Praveen K Gupta, R B Bind, Sameer S Walunj & Mohini Saini

 

 

 

 

 

Sequencing and comparative analysis of hexon gene of fowl adenovirus 4 of Indian origin

 

367

 

        IPC Code: Int. Cl.7 C12N15/09

 

 

        S Barua, B Mondal, A Sanyal, D Hemadri, S K Bandyopadhyay & A Rai

 

 

 

 

 

Typing of bluetongue virus serotype 1 and 23 by RT-PCR

373

 

        IPC Code: Int. Cl.7 C12N15/10, 15/46

 

 

      Swati Dahiya, G Prasad, Minakshi & Ramesh C Kovi

 

 

 

 

 

Generation of a minigenome with non-coding sequences of infectious pancreatic necrosis virus

 

378

 

        IPC Code: Int. Cl.7 C12N15/10, 15/51

 

 

        K Riji John, Siba K Samal & Abdul S Yunus

 

 

 

 

 

Identification of an antiviral principle in Spirulina platensis against Bombyx mori Nuclear Polyhedrosis Virus (BmNPV)

 

384

 

        IPC Code: Int. Cl.7 C12N15/10; G01N33/53

 

 

        S Mahesh Babu, G Gopalaswamy & N Chandramohan

 

 

 

 

 

Genetic analysis of silkworms (Bombyx mori) through RAPD markers

389

 

        IPC Code: Int. Cl.7 C12N15/10

 

 

        P P Srivastava, K Vijayan, A K Awasthi, P K Kar, K Thangavelu & B Saratchandra

 

 

 

 

 

Micropropagation of Adhatoda vasica Nees–A woody medicinal plant by shoot tip culture

 

396

 

        IPC Code: Int. Cl.7 A01H4/00, 5/04

 

 

        Sangeeta Nath & Alak K Buragohain

 

 

 

 

 

Plant regeneration from tissue culture of Chloris virgata: A salt-tolerant desert grass

400

 

        IPC Code: Int. Cl.7 A01H4/00, 5/10

 

 

        Harchand R Dagla & N S Shekhawat

 

 

 

 

 

In vitro propagation and microrhizome induction in Kaempferia galanga Linn. and
K. rotunda Linn.

 

404

 

        IPC Code: Int. Cl.7 A01H4/00, 5/04

 

 

        P Chirangini, S K Sinha & G J Sharma

 

 

 

 

 

Rooting and hardening of in vitro plantlets of Garcinia indica Chois.

409

 

        IPC Code: Int. Cl.7 A01H4/00, 5/10

 

 

        Meera M Chabukswar & Manjushri A Deodhar

 

 

 

Somatic embryogenesis and plantlet regeneration from cotyledon and leaf explants of Solanum surattense

 

414

 

        IPC Code: Int. Cl.7 A01H4/00, 5/10, 5/12

 

 

        N Rama Swamy, T Ugandhar, M Praveen, P Venkataiah, M Rambabu

M Upender & K Subhash

 

 

Short Communications

 

 

 

 

 

Microbial Bioagents: Economic multiplication and management of fungal-nematode complex on cumin

 

419

 

        IPC Code: Int. Cl.7 A01N63/04

 

 

        Nidhi Sharma & P C Trivedi

 

 

 

Electrophoretic studies in induced mutants of diploid mulberry genotype S13

422

 

        IPC Code: Int. Cl.7 C25B7/00; C07K1/26

 

 

        P M Muniswamy Reddy & Munirajappa

 

 

 

 

 

Instructions to Contributors

425

 

 

 


 

AUTHOR INDEX

 

Altaf, M                      342

Awasthi, A K               389

Babu, S M                 384

Bandyopadhyay S K     367

Barua, S                      367

Bhadrayya, K              342

Bind, R B                    363

Buragohain, A K          396

Chandramohan, N        384

Chirangini, P                404  

Chabukswar, M M        409

Dagla, H R                   400

Dahiya, S                     373

Deodhar, M A              409

Devi, J                         307

Dhaneshwar, S S          316

Hemadri, D                  367

Gairola, N                    316

Gopalaswamy,G           384

Gupta, P K                  363

John, R K                    378

Joshi, S N                    316

Kar, P K                      389

Kesri, A                       358

Ko, J M                       307

Kovi, R C                    373

Latha, B V                   353

Mahalakshmi, A           323

Manja, K S                  353

Minakshi                      373

Mittal, B                      358

Mondal, B                   367

Mukherjee, M              358

Munirajappa                 422

Murali, H S                  353

Nampoothiri, K M        336

Nath, S                        396

Naveena, B J                342

Pandey, A                    336

Patel, A K                    336

Prakasham, R S            347

Prasad, G                     373

Praveen, M                  414

Rai, A                          367

Rambabu, M                414

Ramachandran, S         336

Rama Swamy, N          414

Reddy, G                     342

Reddy, P M M             422

Saini, M                       363

Samal, S K                   378

Sanyal, A                     367

Saratchandra, B            389

Sarma, P N                  347

Seo, B B                      307

Sharma, G N               404

Sharma, N                   419

Shenbagarathai, R        323

Shekhawat, N S           400

Sinha, S K                   404

Sreenivas Rao, R          347

Srivastava, A                358

Srivastava, P P             389

Subba Rao Ch              347

Subhash, K                  414

Sujatha, K                   323

Szakacs, G                  336

Thangavelu, K             389

Trivedi, P C                419

Ugandhar, T                414

Upender, M                 414

Venkataiah, P              414

Vijayan, K                   389

Walunj, S S                 363

Yunus, A S                  378  


 

 

 



 

Indian Journal of Biotechnology

Vol.4, July 2005, pp. 307-315

 

 

FISH and GISH: Modern cytogenetic techniques

J Devi1*, J M Ko2 and B B Seo3

1Department of Plant Breeding and Genetics, Assam Agricultural University, Jorhat 785 013, India

2 National Yeongnam Agricultural Experiment Station, Milyang 627 130, Korea

3 Department of Biology, Kyungpook National University, Taegu 702 701, Korea

Received 19 March 2004; revised 23 July 2004; accepted 5 August 2004

In recent years, advances in the molecular cytogenetic technique of fluorescence in situ hybridization (FISH), which enables the direct chromosomal localization of labelled DNA probes and genomic in situ hybridization (GISH), which determines the inter-species distribution of repeated sequences have enabled a resurgence of cytogenetic analysis in plant genome research and molecular breeding. Practical applications of these techniques in chromosome mapping, genome analysis, determination of phylogenetic relationship, detection of chromosomal aberrations and alien chromatin in plant breeding programmes, study of chromosome organization at interphase nuclei and analysis of somaclonal variations in tissue culture have been presented.

Keywords : FISH, GISH, in situ hybridization, rDNA, physical map, probe labelling

IPC Code: Int. Cl.7 A01H1/00

 

 

Indian Journal of Biotechnology

Vol. 4, July 2005, pp. 316-322

 

 

 

RNA Interference: A revolution in drug development

Neha Gairola1, Sonali N Joshi2 and Suneela S Dhaneshwar1*

1Bharati Vidyapeeth Deemed University, Poona College of Pharmacy, Erandwane, Pune 411 038, India

2Department of Microbiology, Lady Amritabai Daga College, Nagpur 440 010, India

Received 18 May 2004; revised 17 August 2004; accepted 28 August 2004

RNA interference (RNAi) or gene silencing technology is a phenomenon by which double stranded RNAs elicit degradation of a target mRNA containing homologous sequence. It is essentially a new incarnation of well-established antisense principle. This technology enables the researchers to trigger post-transcriptional gene silencing in vivo. It is a robust method for lowering specific protein levels, compared to traditional techniques such as antisense, ribozymes or microinjection of function-blocking antibodies. RNAi offers a powerful tool for ascribing functions to genes while its application to in vivo models of disease opens up tremendous opportunities to develop a novel generation of oligonucleotide-based drugs, thus offering an enormous potential of being developed as a therapeutic modality.

Keywords: antisense technology, RNA interference (RNAi), gene silencing, short interfering RNAs (siRNA)

IPC Code: Int. Cl.7 A61K31/713; 48/00

 

Indian Journal of Biotechnology

Vol. 4, July 2005, pp. 323-335

 

 

Analysis of PCR products for PHB production in indigenous
Pseudomonas sp. LDC-5

K Sujatha, R Shenbagarathai* and A Mahalakshmi

PG Department of Zoology and Research Centre, Lady Doak College, Madurai 625 002, India

Received 18 November 2003; revised 12 August 2004; accepted 30 August 2004

Poly-b-hydroxybutyric acid (PHB) and similar bacterial polyesters are promising candidates for the development of environment-friendly, totally biodegradable plastics. The cost of these biopolymers is 25% more than the synthetic polymers that prevents their usage in wider range of applications. In order to reduce the cost, much effort has been made to screen the promising indigenous PHB producing strain in the present study. As a first step, among thirty scl (Short-Chain-Length) positive strains screened, the most promising mcl (Medium Chain Length) PHA positive indigenous isolate, Pseudomonas LDC-5 was selected for further characterization. The nucleotide sequence analysis of the specific PCR product revealed open-reading frames probably relevant for PHA biosynthesis. The similarity search for nucleotide sequence exhibited 92% homology with Pseudomonas sp. The amino acid sequences of the putative proteins deduced from these genes indicate that they encode a PHA synthase, which exhibited 96% amino acid identity with PHA synthase from Pseudomonas putida. Sequence alignment of the partial sequence of PHA synthase genes and putative protein showed conserved signatures. Phylogenetic analysis further places the origin of indigenous isolate closer to P. putida. The partial sequence of PHA synthase was submitted in EMBL and obtained the accession number AJ586810. FT-IR spectral analysis confirmed the presence of strong characteristic ester carbonyl band at 1733cm-1. The identification of this novel biopolymer producing strain reveals a capability for the synthesis of technically interesting biopolymers in future.

Keywords: Poly-b-Hydroxy butyrate, Cloning, Colony PCR, Pseudomonas sp., FT-IR analysis, phb Operon

        IPC Code: Int.Cl.7: C08G63/00; C12N15/10; C12R1:38

 

 

Indian Journal of Biotechnology

Vol. 4, July 2005, pp. 336-341

 

 

 

Partial purification and characterization of a-amylase produced by Aspergillus oryzae using spent-brewing grains

Anil Kumar Patel1, K Madhavan Nampoothiri1, Sumitra Ramachandran1

George Szakacs2 and Ashok Pandey1*

1Biotechnology Division, Regional Research Laboratory (CSIR), Thiruvananthapuram 695 019, India

2Department of Agricultural Chemical Technology, Technical University of Budapest, 1111 Budapest, Hungary

Received 23 December 2003; revised 13 July 2004; accepted 25 July 2004

Solid-state fermentation (SSF) was carried out to produce a-amylase from Aspergillus oryzae (IFO 30103) using spent brewing grains (SBG) as substrate. A maximum of 11296 U/gds amylase activity was obtained after 48 h of fermentation. The extracted enzyme was subjected to partial purification by ammonium sulphate fractionation. Maximum specific activity was obtained with 40-70% fraction. SDS-PAGE of the corresponding sample revealed an approximate 66 kDa band, which was confirmed by activity staining as a-amylase. It was optimally active at pH 5 and 50°C by using 1% starch as substrate concentration. The partially purified a-amylase loses activity rapidly above 50oC but it can be retained in the presence of Ca2+. Presence of Mn2+ and Fe2+ enhanced the enzyme activity and is almost doubled in presence of Mn2+. However, in the presence of Hg2+ and Cu2+, the activity is reduced. Hg2+ reduced the enzyme activity by half.

Keywords: a-Amylase, Aspergillus oryzae, solid-state fermentation, spent brewing grains

            IPC Code: Int. Cl.7 A01N63/02; C12N9/30; C12R1: 69

 

 

 

Indian Journal of Biotechnology

Vol. 4, July 2005, pp. 342-346

 

 

 

Screening and interaction effects of physical parameters, total N content and buffer on L(+) lactic acid production in SSF by Lactobacillus amylophilus
GV6 using Taguchi designs

B J Naveena1, Md Altaf1, K Bhadrayya2 and Gopal Reddy1*

1Department of Microbiology, Osmania University, Hyderabad 500 007, India

2Swaroop Tech Consultancy, Secunderabad 500 003, India

Received 28 October 2003; revised 11 September 2004; accepted 22 September 2004

Lactobacillus amylophilus GV6 was studied for L(+) lactic acid production in solid state fermentation. The physical parameters, such as moisture, temperature, pH, inoculum size, incubation period, total N content and buffering agent (CaCO3), were studied using the Taguchi statistical methods to observe the interaction effects and identify the vital factors governing the process of fermentation beneficial in further improvement of lactic acid production. Moisture is identified to be the important physical parameter, showing maximum impact at 83%, along with temperature 37oC, pH 6.5, inoculum size 9 mL, incubation period 9 days, total N content 2% and CaCo3 1.5 g/10 grams of substrate (wheat bran).

Keywords: lactic acid, Lactobacillus amylophilus, solid state fermentation, Taguchi method, wheat bran

            IPC Code: Int. Cl.7 A01N63/02; C12N9/00; C12R1: 225

 

 

Indian Journal of Biotechnology

Vol. 4, July 2005, pp. 347-352

 

 

 

Cyclodextrin glycosyl transferases from Bacillus circulance and Bacillus sp.

R S Prakasham*, R Sreenivas Rao, Ch Subba Rao and P N Sarma

Biochemical and Environmental Engineering Centre, Indian Institute of Chemical Technology, Hyderabad 500 007, India

Received 26 April 2004; revised 20 July 2004; accepted 5 August 2004

Cyclodextrin glycosyl transferase production and its activity was evaluated using locally isolated Bacillus sp. and Bacillus circulance. The enzyme production and its induction processes are dependent on the type of the organism. Isolated Bacillus sp. possesses higher enzyme productivity and early induction to that of B. circulance. The enzyme synthesis in both these strains was observed to be dependent on the carbon source and inoculum level. Activity related experimental results denoted that the properties of the enzyme vary with the source of the organism with respect to hydrogen ion concentration, temperature profile and presence of the ionic species. Overall, the data indicate that both organisms differ in CGTase production and isolated Bacillus sp. possesses higher potential as enzyme producer to that of B. circulance.

Keywords: Bacillus sp., Bacillus circulance, CGTase production, enzyme

            IPC Code: Int. Cl.7 A01N63/02; C12N9/24; C12R1:07, 1:09

 

 

 

Indian Journal of Biotechnology

Vol. 4, July 2005, pp. 353-357

 

 

 

Influence of growth factors on carotenoid pigmentation of
Rhodotorula glutinis DFR-PDY from natural source

B V Latha, K Jeevaratnam*, H S Murali and K S Manja

Defence Food Research Laboratory, Siddarthanagar, Mysore 570 011, India

Received 31 July 2003; revised 2 July 2004; accepted 15 July 2004

A red yeast (Rhodotorula glutinis DFR-PDY) strain, isolated from a contaminated Potato Dextrose Agar (PDA) plate, was evaluated for its growth and pigment production. The yeast was able to grow and produce carotenoid pigments over a wide range of pH from 2.5 to 9.5 (optimum pH 5.5) and at ambient temperature of 29-32oC in a modified Czapek Dox broth after 12 days of incubation. However, light was not a limiting factor for the growth and pigmentation of the red yeast. Among the various nitrogen and carbon sources examined, maximum growth and pigmentation was observed in sodium nitrate as nitrogen source and glucose and fructose as carbon sources. Thin layer chromatography of the extracted carotenoid pigments revealed the presence of four colour components.

Keywords: carotenoids, fermentation, Rhodotorula glutinis

            IPC code: Int. Cl.7 A01N63/02; B01D15/08; C12P23/00; CR1:645

 

 

Indian Journal of Biotechnology

Vol. 4, July 2005, pp. 358-362

 

 

Analysis of VNTR loci, ApoB 3˘ HVR and D1S80 in North Indians

Monisha Mukherjee, Anvesha Srivastava, Akanchha Kesari and Balraj Mittal*

Department of Medical Genetics, Sanjay Gandhi Post Graduate Institute of Medical Sciences, Lucknow 226 014, India

Received 13 January 2004; revosed 23 July 2004; accepted 10 August 2004

Polymorphic markers like VNTRs at Apolipoprotein B 3˘ and locus D1S80 hypervariable region have been used extensively for population studies through out the world. In the present study, the polymorphism data in North Indian population at these VNTR loci was reported. The allele distributions and their genotype frequencies at the VNTR loci, Apo B and D1S80 were reported in 86 (172 chromosomes) and 75 (150 chromosomes) unrelated normal individuals, respectively. Genomic DNA was extracted from blood samples and amplified by polymerase chain reaction. The respective alleles and their sizes were determined, 19 and 24 different alleles making up 51 and 50 genotypes of Apo B and D1S80 respectively were identified in the North Indian population. As per c2 analysis, the allele and genotype frequencies for both VNTRs were in Hardy-Weinberg equilibrium. The most frequent allele of Apo B (allele 6) corresponded to 40 repeats, and D1S80 (allele 12) to 28 repeats. The frequencies were 0.087 and 0.17 and observed heterozygosities were 55 and 57% for Apo B and D1S80, respectively. This information may have implications in disease diagnostics, forensics, paternity analysis, and for ruling out maternal contamination in fetal samples during prenatal diagnosis of genetic disorders.

Key words: Apo B, D1S80, hypervariable region, maternal contamination, prenatal diagnosis, variable number of tandem repeats

IPC Code: Int. Cl.7 C12N15/10

 

 

 

Indian Journal of Biotechnology

Vol. 4, July 2005, pp. 363-366

 

Cloning, characterization and expression of bovine (Bos indicus)
 tumour necrosis factor-a

Praveen K Gupta1*, R B Bind1, Sameer S Walunj1 and Mohini Saini2

1National Biotechnology Centre, 2Division of Biochemistry and Food Sciences, Indian Veterinary Research Institute
Izatnagar 23 122, India

Received 23 January 2004; revised 2 August 2004; accepted 16 August 2004

The gene for tumour necrosis factor-alpha (TNF-a) was amplified from cDNA pool prepared from LPS-stimulated bovine peripheral blood mononuclear cells isolated from Indian cattle. The amplified TNF-a gene was cloned and nucleotide sequences were determined. Homology comparison of nucleotide and predicted amino acid sequences revealed similarity at nucleotide and amino acid level in both exotic cattle and goat. The coding sequence of mature TNF-a (without its signal sequence and transmembrane anchor) was expressed as fusion protein with N-terminal polyhistidine using prokaryotic expression vector. The expressed protein was present as insoluble inclusion bodies. The recombinant protein was solubilized with urea and purified using Ni-agarose affinity chromatography. The purified recombinant TNF-a was characterized in SDS-PAGE and in western blotting.

Keywords: Bos indicus, cDNA sequence, expression, TNF-a, prokaryotic, recombinant

            IPC code: Int. Cl.7 C12N15/10, 15/28

 

 

Indian Journal of Biotechnology

Vol 4, July 2005, pp 367-372

 

 

 

Sequencing and comparative analysis of hexon gene of fowl adenovirus 4 of Indian origin

S Barua1, B Mondal2, A Sanyal2, D Hemadri2, S K Bandyopadhyay and A Rai*

Division of Veterinary Biotechnology, Indian Veterinary Research Institute, Izatnagar 243 122, India

1Central Institute of Research on Goats, Makhdoom, Farah, Mathura 281 122, India

2IVRI, Mukteswar, Kumaon 263 138, India

Received28 May 2004; revised 21 July 2004; accepted 5 August 2004

The sequencing of the recombinant plasmid containing 2916 bp hexon gene insert of fowl adeno virus 4 (FAV4) of Indian origin revealed the complete nucleotide sequence of the FAV4 hexon gene. The sequence analysis revealed an open reading frame of 2814 bp coding for a 937 amino acids long polypeptide with a molecular mass of 106.04 kDa. This sequence was larger than the hexon coding regions of FAV10, HAV40, HAV41, MAV1 and BAV3, while it was smaller than the hexon in HAV2 and HAV5. The Kozak sequence was found at nucleotide position 78, which was similar in FAV10 hexon gene. The codon bias revealed a strong preference for C and to a lesser extent G in the third base position. The splice acceptor sequence in FAV4Ind was located 12 bp upstream of the hexon initiation site of the translation codon, which indicated that FAV4 hexon could be a late gene product. A high degree of amino acid variation (9.6%) was observed between FAV4Ind and FAV10, as compared to the 1.5% variation between FAV4Ind and FAV4KR95. The L1 loop of the hexon gene showed the maximum sequence variation among FAV serotypes, thus it could be used for designing serotype specific PCR primers. The 11 unique amino acid substitutions in FAV4Ind hexon could play a significant role in determining its type specificity.

Key words: fowl adenovirus 4, hexon gene, sequencing, virus

            IPC Code: Int. Cl.7 C12N15/09

 

 

Indian Journal of Biotechnology

Vol 4, July 2005, pp 373-377

 

 

Typing of bluetongue virus serotype 1 and 23 by RT-PCR

Swati Dahiya, G Prasad*, Minakshi and Ramesh C Kovi

Department of Animal Biotechnology, CCS Haryana Agricultural University, Hisar 125 004, India

Received 5 March 2004; revised 26 July 2004; accepted 10 August 2004

Two primer pairs were evaluated for amplification of unique regions on the serotype-specific segment 2 (VP2 gene) of bluetongue virus (BTV) by reverse transcription-polymerase chain reaction (RT-PCR). A new set of primer pair [forward primer (FP): 1240-1271 bp and reverse primer (RP): 1844-1813 bp] specific to VP2 gene of BTV serotype 1 (BTV-1) was designed. This primer pair successfully amplified the cell-culture-grown six BTV-1 Indian isolates from different geographical regions, yielding an expected PCR product of 604 bp. However, the primer pair failed to amplify Indian isolates of BTV-18 and BTV-23. Similarly, the primer pair for BTV-23 serotype (FP: 604-623 bp; RP: 1262-1243 bp) amplified 658 bp amplicon with Indian isolates of only BTV-23 and not with the isolates of BTV-1 and BTV-18 serotypes. These VP2 gene based serotype specific primers when aligned with other BTV serotype sequences available in the GenBank revealed no significant sequence identity except with the serotypes for which they were designed. This strongly suggests that the serotype specific primers have potential application for detection of BTV-1 and BTV-23 by using rapid, reliable and specific RT-PCR assay.

Keywords: Bluetongue virus, RT-PCR, serotyping, VP2 gene

            IPC Code: Int. Cl.7 C12N15/10; 15.46

 

 

 

Indian Journal of Biotechnology

Vol. 4, July 2005, pp. 378-383

 

Generation of a minigenome with non-coding sequences of infectious pancreatic necrosis virus

K Riji John1*, Siba K Samal and Abdul S Yunus

Department of Veterinary Medicine, Virginia-Maryland Regional College of Veterinary medicine,
University of Maryland, MD 20742, USA

Received 11 November 2003; revised 13 July 2004; accepted 28 July 2004

Infectious pancreatic necrosis virus (IPNV) is an important aquatic pathogen causing a highly devastating disease in salmonids. The virus has been isolated from over 50 species across the world. For combating the disease, vaccines have been developed by different recombinant DNA technologies. Production of live virus vaccines with defined attenuations requires reverse genetics system and minigenome synthesis to study the attenuation and virus production in vitro systems. Towards this objective, the two open reading frames of the IPNV (West Buxton strain) were genetically engineered to replace them with bacterial chloramphenicol acetyl transferase (CAT) reporter gene while retaining the non-coding regions (NCR). The minigenome of IPNV without the coding regions was generated using a modified pUC 19 plasmid and was checked for the nucleotide correctness by dideoxy chain termination method. Expression of the reporter gene was verified after transfection studies in susceptible cell line. The synthesised minigenome is useful in carrying out a number of studies in the reverse genetics of IPNV.

Keywords: IPNV, minigenome, reverse genetics, virus vaccine

            IPC Code: Int. Cl.7 C12N15/10, 15/51

 

 

 

Indian Journal of Biotechnology

Vol. 4 July, 2005, pp. 384-388

 

Identification of an antiviral principle in Spirulina platensis against Bombyx mori Nuclear Polyhedrosis Virus (BmNPV)

S Mahesh Babu1, G Gopalaswamy1* and N Chandramohan2

1Department of Biotechnology and 2 Department of Sericulture, Tamil Nadu Agricultural University, Coimbatore 641 003, India

Received 28 July 2003; revised 1 April 2004; accepted 20 August 2004

Virus diseases in Bombyx mori, particularly the Nuclear Polyhedrosis Virus (NPV) account for 70-80% of the total loss and thus pose a major problem in sericulture. The NPV is characterized by having distinctive inclusion bodies either polyhedral or capsular in size, in which several virus particules are embedded. Spirulina has been found to inhibit the viral replication and strengthen both the cellular and humoral arms of the immune system. In the present study, S. platensis at various concentrations. viz, 2, 4, 6, 8, 10, 12, 14, 16, 18 and 20% were given to the third instar silkworm larvae by leaf dip method, as a feed supplement along with BmNPV. A separate control (larvae fed with BmNPV alone) was maintained. The results showed that the silkworm fed with S. platensis at 10% concentration showed 90% resistance to BmNPV. This resistance was found to be due to an antiviral protein in fraction IV (60-80%) of molecular weight 29 kDa. The protein fraction was purified by HPLC and collected separately. An antibody was raised against this protein in rabbit. Western blotting, along with other serological techniques like agglutination test and precipitation test were conducted to confirm the result.

Keywords: Spirulina platensis, Bombyx mori, antiviral, BmNPV, immunological tests

IPC Code: Int. Cl.7 C12N15/10, G01N33/53

 

 

 

Indian Journal of Biotechnology

Vol. 4, July 2005, pp. 389-395

 

 

Genetic analysis of silkworms (Bombyx mori) through RAPD markers

P P Srivastava1*, K Vijayan1, A K Awasthi1, P K Kar2, K Thangavelu2 and B Saratchandra1

1SeriBiotech Research Laboratory, Central Silk Board, Carmelram Post, Kodathi, Bangalore 560 035, India

2Central Sericultural Germplasm Resources Centre, Hosur 635 109, India

Received 19 August 2003; revised 17 May 2004; accepted 2 June 2004

Mulberry silkworm, Bombyx mori L. is the most important insect being used for commercial extraction of silk in sericulture industry. To enhance the productivity and quality of silk fibers, many attempts are being made to improve the silkworm stocks through genetic manipulation. For that knowledge on the genetic diversity of the parental stocks is essential for selection and conservation of these precious materials. Keeping this in view, genetic diversity among twenty silkworm stocks differing in their yield potential and nature of voltinism was estimated through PCR amplification of the genomic DNA with 10 random amplified polymorphic DNA (RAPD) primers. A total of 68 bands were generated. Of which 61 were polymorphic (90%). The pair-wise genetic distance estimated from these bands varied from 0.058 to 0.513 with an average distance of 0.246. The cluster analysis using unweighted pair group using arithmetic average (UPGMA) grouped the silkworms into six groups and one isolate. Spatial distribution of the silkworms on two-dimensional figures using ALSCAL multidimensional scaling has broadly discriminated the multivoltine from the bivoltine silkworms. Certain silkworm stocks like Kalimpong-A, C’nichi, Nistari (P) and Mysore Princess were having higher genetic distance from others and, thus, could be used for heterosis breeding and also in breeding programs aimed at introgressing hardy genes into the bivoltine high yielding stocks.

Keywords: Bombyx mori, genetic variability, RAPD markers

            IPC Code: Int.Cl.7: C12N15/10

 

 

Indian Journal of Biotechnology

Vol. 4, July 2005, pp. 396-399

 

 

Micropropagation of Adhatoda vasica Nees–A woody medicinal plant
by shoot tip culture

Sangeeta Nath and Alak K Buragohain*

Department of Molecular Biology and Biotechnology, Tezpur University, Napaam, Tezpur 784 028, India

Received 17 November 2003; revised 19 August 2004; accepted 2 September 2004

Exudation was overcome during shoot tip (1 cm) culture of Adhatoda vasica by modifying the pH of the media and by lowering the concentrations of NH4+ and K+ ions. Though shoot formation occurred in media supplemented with BA
(2 mg L-1), 96.67% of the shoots developed callus at the cut basal end and the explants turned brown and necrotic due to the phenolic exudates released into the medium. Development of callus tissue and browning was eliminated by culturing the shoot tip explants on primary MS medium supplemented with thiadiazuron (0.30 mg L-1) and coconut milk (15%) which also promoted development of shoots but with stunted curled leaves. Shoots (5.2) with healthy leaves were obtained when these shoots were transferred to a secondary MS medium supplemented with 0.5 mgL-1 BA. Incorporation of coconut milk (15%) in the secondary MS medium had a growth promontory effect. High frequency rooting (9.33) was recorded in MS basal medium. The rooted plantlets were hardened and established at 85% rate in pots.

Keywords: thiadiazuron; Adhatoda vasica; phenolic exudates; shoot tip; micropropagation

            IPC Code: Int. Cl.7 A01H 4/00, 5.04

 

 

 

Indian Journal of Biotechnology

Vol. 4, July 2005, pp. 400-403

 

 

Plant regeneration from tissue culture of Chloris virgata:
A salt-tolerant desert grass

Harchand R Dagla and N S Shekhawat*

Biotechnology Unit, Department of Botany, Jai Narain Vyas University, Jodhpur 342 001, India

Received 15 April 2004; revised 13 July 2004; accepted 25 July 2004

A tissue culture protocol for regeneration of salt-tolerant grass Chloris virgata Sw. is reported. Scutellum and axis of activated embryo produced callus on Murashige & Skoog (MS, 1962) medium with 2.0-45.0 µM of 2,
4-dichlorophenoxyaceticacid (2,4-D). The explants produced two types of callus. Type-I produced on MS + 2.0-9.0 µM 2,
4-D was amorphous, translucent and white; while Type-II produced on 22.0-45.0 µM 2,4-D was creamy, compact and globular. The latter exhibited competence to regenerate into plants. These cultures were amplified on MS + 22.0 µM 2,4-D. The regenerative cultures, on transfer to full, half and one-fourth strengths of growth regulator-free MS or full MS + 2.0 to 8.0 µM benzyladenine (BAP) differentiated into plantlets. The highest regeneration potential (6-7 plantlets per inoculation) was achieved on full-strength growth regulator-free MS medium with 50 mg L-1 ascorbic acid and 25 mg L-1 each of adenine sulphate, arginine and citric acid. The tissue culture raised plants were hardened in a greenhouse with controlled environmental conditions. Ninety per cent of the regenerants survived during acclimatization. A total of 504 acclimatized plants were transferred to polybags and in field. Of these 80% survived in the polybags and 70% in the field.

Keywords: Chloris virgata, desert grass, salt tolerant, plantlet regeneration, scutellum.

            IPC Code: Int. Cl.7 A01H4/00, 5/10

 

 

Indian Journal of Biotechnology

Vol. 4, July 2005, pp. 404-408

 

 

In vitro propagation and microrhizome induction in Kaempferia galanga Linn. and K. rotunda Linn.

P Chirangini, S K Sinha1 and G J Sharma*

Department of Life Sciences, Manipur University, Imphal 795 003, India

Received 2 April 2004; revised 11 August 2004; accepted 25 August 2004

Rhizomatous buds of Kaempferia galanga and K. rotunda induced microshoots when cultured on Murashige and Skoog (MS) medium supplemented with plant growth regulators. Multiple shoots were induced on MS medium containing 5.70 mM IAA alone and a combination of 0.57 mM IAA plus 4.65 mM Kn in case of K. galanga. Whereas, the medium supplemented with 2.69 mM NAA plus 2.22 mM BAP was the best for K. rotunda. Further, subculture of the microshoots gave more multiple shoots (13) on medium containing 4.44 mM BAP in K. galanga and (9) on the 2.69 mM NAA and
2.22
mM BAP in K. rotunda. Induction of callus was observed in K. galanga on the medium supplemented with 2.85 mM IAA. Microshoots were regenerated from the callus on 2.69 mM NAA and 2.22 mM BAP enriched medium. Microrhizome formation was observed within one month of incubation of microshoot cultures (~4 month old, following 4 passages) in the medium supplemented with 6-9% sucrose with either 22.2 mM BAP or 23.25 mM Kn. These microrhizomes produced shoots when transferred to fresh microshoot induction medium within 2-3 weeks of incubation. The microshoots produced roots irrespective of their method of regeneration. Plantlets transplanted to pots grew to mature plants after 3 months of transfer and showed 80-90% survival.

Keywords: 6-benzylaminopurine (BAP), indole-3-acetic acid (IAA), Kaempferia, kinetin (Kn), microrhizome, α-naphthalene acetic acid (NAA)

            IPC Code: Int. Cl.7 A01H4/00, 5/04

 

 

Indian Journal of Biotechnology

Vol. 4, July 2005, pp. 409-413

 

 

Rooting and hardening of in vitro plantlets of Garcinia indica Chois

Meera M Chabukswar and Manjushri A Deodhar*

Kelkar Education Trust’s V G Vaze College of Arts, Science and Commerce, Mithagar Road, Mulund (E) Mumbai 400 081, India

Received 5 December 2003; revised 31 August 2004, accepted 15 September 2004

In the present paper, a procedure for rooting and acclimatization of in vitro plantlets raised from immature seed cultures of Garcinia indica is described. Pulse treatment of 9800µM IBA was preferred for rooting of in vitro shoots. Nearly 100% rooting (in vitro) was obtained with half strength MS with 2% sucrose and 0.6 % agar. Rooted plantlets of about 2-4 cm height (2-3 nodes) were subsequently transferred to different media for acclimatization: 1. Cocopeat + Sand + Soil (1:2:1); 2. Cocopeat + Sand (1:1); 3. Cocopeat. Plants grown in cocopeat showed 76% of survival followed by cocopeat and sand (1:1) with 75% of survival. The in vitro elongated shoots could also be rooted ex vitro by treating with synthetic hormones like Keradix, Rootex-1, and Rootex-3. However, the success of rooting was low (20 to 55 %). The hardened plantlets were successfully acclimatized in the greenhouse and transferred to open field conditions.

Keywords: Garcinia indica, micropropagation, rooting, hardening

            IPC code: Int. Cl.7 A01H4/00, 5/10

 

 

Indian Journal of Biotechnology

Vol. 4, July 2005, pp. 414-418

 

 

Somatic embryogenesis and plantlet regeneration from cotyledon and leaf explants of Solanum surattense

N Rama Swamy*, T Ugandhar, M Praveen, P Venkataiah, M Rambabu, M Upender and K Subhash

Plant Biotechnology Group, Department of Botany, Kakatiya University, Warangal 506 009, India

Received 14 May 2004; revised 22 July 2004; accepted 5 August 2004

Somatic embryogenesis and plant regeneration from the cotyledon and leaf explants of Indian Solanum (Solanum surattense Burm. f.), a medicinally important plant, is reported. Embryogenic callus was induced from cotyledon and leaf explants on Murashige and Skoog’s medium fortified with 0.5-8.0 mg/L α-naphthalene acetic acid (NAA) + 0.5 mg/L N6-benzylaminopurine (BAP). High frequency of somatic embryo formation was found at 6.0 mg/L NAA + 0.5 mg/L BAP and 4.0 mg/L NAA+ 0.5 mg/L BAP in cotyledon and leaf explants, respectively. Secondary somatic embryogenesis was also observed when primary somatic embryos were subcultured on the same somatic embryo induction medium. Well-developed cotyledonary stage embryos were germinated on MS medium supplemented with 0.5 mg/L indole-3-acetic acid (IAA) +1.0-8.0 mg/L BAP. Maximum percentage (71.2%) of somatic embryo germination and plantlet formation was found at 0.5 mg/L IAA+2.0 mg/L BAP, but they didn’t germinate on ˝ MSO and MSO media. The post transplantation survival rate of plants was 65-70%. Plants and flowers formed were morphologically similar to mother plants. The present protocol can be used for genetic transformation experiments in S. surattense.

Keywords: S. surattense, Indian solanum, somatic embryos, regeneration

IPC code: Int. Cl.7 A01H 4/00, 5/10, 5/12

 

 

Indian Journal of Biotechnology

Vol. 4, July 2005, pp. 419-421

 

 

Microbial Bioagents: Economic multiplication and management of fungal-nematode comple´ on cumin

Nidhi Sharma* and P C Trivedi

Department of Botany, University of Rajasthan, Jaipur 302 004, India

Received 21 October 2003; revised 12 August 2004; accepted 30 August 2004

Mass-scale multiplication was done on cheaper substrates for application of bioagents in the management of plant-parasitic nematodes and fungal pathogens. The bioagents were isolated from the local field soils. Out of the 13 isolated fungi, most of the isolates of Trichoderma spp. that were found antagonistic to Fusarium o´ysporum f. sp. cumini in dual culture technique, were mass multiplied on cheaper agrowastes. Suitability of 6 substrates was screened and tea waste was found to be best followed by wheat bran and sorghum straw. Trichoderma harzianum (T5) had the ma´imum spore load per gram (SLPG) value on tea waste followed by T. hamatum (T16) on wheat bran. Three isolates of bacteria viz. Bacillus subtitlis, Pseudomonas fluorescens and Rhizobium spp. were multiplied on nutrient broth, King’s B broth and yeast e´tract mannitol broth, respectively.

Keywords:           biocontrol, Meloidogyne incognita, Fusarium o´ysporum f. sp. cumini, Trichoderma harzianum, T. viride, T. hamatum, Bacillus subtilis, Pseudomonas fluorescens

IPC Code: Int. Cl.7 A01N 63/04

 

 

Indian Journal of Biotechnology

Vol. 4, July 2005, pp. 422-423

 

 

Electrophoretic leaf protein profiles of Morus alba variety S13, irradiated with different dosages of gamma radiation, revealed proteins (polypeptides) of both high and low molecular weights; 44 kDa was the major protein component. Gamma irradiation influenced the quantitative differences in the minor components and low molecular weight proteins present in trace amounts. Significant decrease in the quantity of 55 kDa protein was observed with the increase in dosage from 7 kR to 10 kR. thus, necessitating to limit the dosage of gamma irradiation to 6 kR.

Keywords: leaf protein profiles, mulberry, silver staining

IPC Code: Int. Cl.7 C25 B7/00: C07K1/26

 

 


INDIAN JOURNAL OF BIOTECHNOLOGY

 

Instructions to Contributors

 


The Indian Journal of Biotechnology, a quarterly journal, publishes original research papers, reviews, digests (biotechnology highlights), news-scan, etc. The journal covers papers on Biotechnology in the following main areas: (i) Agriculture; (ii) Animal husbandry; (iii) Environment; (iv) Industry; (v) Microbiology; (vi) Medicine; (vii) Bio-informatics; and (viii) Socio-legal and ethical aspects.

Indian Journal of Biotechnology invites original research and review manuscripts not submitted for publication elsewhere. It is mandatory on the part of the corresponding author to furnish the following certificate at the time of submission of the manuscript:

This is to certify that the reported work in the paper entitled "                " submitted for publication is an original one and has not been submitted for publication elsewhere. I/we further certify that proper citations to the previously reported work have been given and no data/tables/figures have been quoted verbatim from other publications without giving due acknowledgement and without the permission of the author(s). The consent of all the authors of this paper has been obtained for submitting the paper to the "Indian J Biotechnology".

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The copyright of the paper will be transferred from the author to publisher. One original and two copies of the manuscript should be submitted to the editor. The manuscript, after referees’ acceptance, will be sent back to the author(s) along with referees’ comments. For re-submission, two copies of the revised version of the manuscript, and a copy on floppy disk [3.5˘˘ (1.44 MB)] using word processing software such as MS Word (version 6 and onwards), or PDF files (version 4 and onwards), or as an attachment to e-mail should be submitted to the editor.

 

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References—References should be cited in the text by the consecutive numbers of their occurrence; the numbers are to be shown as superscript at the end of the statement related to that particular reference, e.g. It also inhibits the activity of endogenous DNA polymerase of HBV7.

Following the same sequence of the text, the list of references be appended under the References heading. Each reference should provide names and initials of all the authors, giving coma in between the authors and ‘&’ before the last author. In case, the authors are more than five, then use et al after the 5th author. It should be followed by title of the paper, abbreviated title of journal (in italics), volume number, year of publication (within circular bracket), and the starting and closing page numbers. Abbreviated titles should conform to the international guidelines, e.g. The Chemical Abstracts Service Source Index (CASSI) or BIOSIS

The style of references should be:

 

Research Papers

·     Ghosh A C & Basu P S, Extracellular polysaccharide production by Azorhizobium caulinodans from stem nodules of leguminous emergent hydrophyte Aeschynomene aspera, Indian J Exp Biol, 39 (2001) 155-159 [If accepted for publication, give (in press) in place of volume, year and pages].

·     Newell C A, Lowe J M, Merryweather A, Rooke L M & Hamilton W D O, Transformation of sweet potato (Ipomoea batatas (L) Lam) with Agrobactericum tumefaciens and regeneration of plants expressing cowpea trypsin inhibitor and snowdrop lectin, Plant Sci, 107 (1995) 215-227.

·     Hoffman M P, Zalom F G, Smilanick J M, Malyj L D, Kiser J et al, Field evaluation of transgenic tobacco containing genes encoding Bacillus thuringensis d-endotoxin or cowpea trypsin inhibitor: Efficacy against Helicoverpa zea (Lepidoptera: Noctuidae), J Econ Entomol, 85 (1991) 2516-2522


Books & Proceedings of Conferences

·     Truzuki T & Irukayama K, Minamata disease (Elsevier, Amsterdam) 1977, 30-45.

·     Roels O A & Mahadevan S, Vitamin A, in The vitamins: Chemistry, pathology and methods, 2nd edn, vol VI, edited by P Gyorgy & W N Pearson (Academic Press, New York) 1967, 139-210.

·     Allossp P G, Nutt K A, Geijsk R J & Smith G R, Transgenic Sugarcane with increased resistance to canegrub, in Sugarcane pest management in the new millennium, 4th Sugarcane Entomol Workshop, held on 7-10 Feb, 2000 (Int Soc Sugarcane Technol, Khon-khon, Thailand) 2000, 63-67.

·     Chaturvedi H C & Sharma A K, Citrus tissue culture, in Proc Natl Semin Plant Tissue Cult (ICAR, New Delhi) 1988, 36-46.

·     Kapoor B C, 2000. Managing in the face of not-so-developed and organized environment, paper presented in Natl Symp Manag Dev, Institute of Public Administration, Jaipur, India, 21-23 July, 2000.

 

Thesis & Dissertation

·     Chaturvedi H C, In vitro growth and controlled morphogenesis in callus tissue of Rauvolfia serpentina. Ph D Thesis, Agra University, Agra, 1968.

 

Patent

·     Trepaginer J H, New surface finishings and coatings, US Pat 1276323 (to DuPont Inc, USA). 27 June, 2000; Chem Abstr, 49 (2000) 27689.

Manuscript along with referees’ comments will be sent to the author identified for correspondence on the title page of the manuscript. It should be checked carefully and the modified manuscript should be returned within ten days of receipt. No page proofs will be sent to author(s).

 

Reprints—Twenty five reprints will be supplied gratis.