Indian Journal of Biotechnology

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VOLUME 8

CODEN: IJBNAR 8(1) (2009) 1-140

NUMBER 1

JANUARY 2009

ISSN: 0972-5849

 

CONTENTS

 

Reviews

 

Recent advances in medicinal plant biotechnology

9

Mohammad Yaseen Khan, Saleh Aliabbas, Vimal Kumar & Shalini Rajkumar

 

 

 

Characterization of embryonic stem cells: A special focus on farm animals

23

D Kumar, T Anand, M K Singh, M S Chauhan & R S Manik

 

 

 

Papers

 

Genotype analysis and assessment of antigenic sensitivity for recombinant HCV proteins by indigenous SIBA for detection of Hepatitis C Virus infection: A comparison with 3rd EIA and RT-PCR

33

S P D Ponamgi, M Chandra, Y Naresh Kumar, S Rahamathulla, Lakshmi Narasu,
C M Habibullah & M N Khaja

 

 

 

Molecular level studies on multiple antibiotic and serum resistance in UTI pathogens

40

S M Dharmadhikari & S A Peshwe

 

 

 

Structure-function annotation and phylogenetic strategy of nifH domain  of a

cyanobacterium — Chlorogloeopsis sp.

46

P T V Lakshmi, S Uma Maheswari & A Annamalai

 

 

 

Down-regulation of an abiotic stress related Nicotiana benthamiana WRKY transcription factor induces physiological abnormalities

53

K Archana, N Rama, H M Mamrutha & Karaba N Nataraja

 

 

 

Molecular diversity in genus Nicotiana as revealed by randomly amplified polymorphic DNA

61

K Siva Raju, M Sheshumadhav, C Chandrasekhararao & T G K Murthy

 

 

 

Use of RAPD markers for assessment of genetic diversity in sugarcane cultivars

67

P G Kawar, R M Devarumath & Y Nerkar

 

 

 

Enhanced insect resistance to bollworm (Helicoverpa armigera) in cotton containing a synthetic cry1Ab gene

72

Farzane Yazdanpanah, M Tohidfar, M Esna Ashari, B Ghareyazi, M Karimi Jashni & M Mosavi

 

 

 

Agrobacterium-mediated transformation of chickpea using shoot meristem

78

Rekha Singh, N P Singh, Subhojit Datta, Indu Singh Yadav & A P Singh

 

Genetic variation and differentiation in the Stinging catfish, Heteropneustes fossilis (Bloch), populations assessed by heterologous microsatellite DNA markers

85

Shamima Nasren, Mohammad Nazrul Islam, Mohd Golam Quader Khan,
Md Shahidul Islam & Md Samsul Alam

 

 

 

Probable mechanism(s) of antifungal activity of HA-2-91—A tetraene antibiotic

91

S R Naik & T E Gupte

 

 

 

Characterization of functional activity in composted casing amendments used in cultivation of Agaricus bisporus (Lange) Imbach

97

Devendra K Choudhary, Pavan K Agarwal & Bhavdish N Johri

 

 

 

Characterization of thermoalkalophilic xylanase isolated from Enterobacter isolates

110

Anjana Sharma, Rajesh Pujari & Pratibha Patel

 

 

 

Micropropagation of Wrightia tomentosa: Effect of gelling agents, carbon source and vessel type

115

Preeti Joshi, Rohini Trivedi & S D Purohit

 

 

 

Regeneration competence of Tainia latifolia (Lindl.) Benth ex Hook pseudobulb segments: An
in vitro study

121

Sungkumlong & Chitta Ranjan Deb

 

 

 

Regeneration of Biophytum sensitivum (Linn.) DC. through organogenesis and somatic embryogenesis

127

M B Shivanna, M M Vasanthakumari & M C Mangala

 

 

 

Direct organogenesis and somatic embryogenesis in Beloperone plumbaginifolia (Jacq.) Nees

132

M C Shameer, V P Saeeda, P V Madhusoodanan & Sailas Benjamin

 

 

 

Instructions to Contributors

137

 

 

 

AUTHOR INDEX

 



 

 

 

 

Indian Journal of Biotechnology

Vol 8, January 2009, pp 9-22

 

 

Recent advances in medicinal plant biotechnology

 

Mohammad Yaseen Khan1, Saleh Aliabbas2, Vimal Kumar1 and Shalini Rajkumar2*

 

1Department of Phytopharmaceuticals and Natural Products, Institute of Pharmacy

2Department of Biochemistry and Biotechnology, Institute of Science,

Nirma University of Science and Technology, Sarkhej-Gandhinagar Highway, Ahmedabad 382 481, India

 

Received 18 July 2007; revised 19 May 2008; accepted 10 August 2008

Medicinal plants are the most important source of life saving drugs for the majority of the world’s population. Plant secondary metabolites are economically important as drugs, fragrances, pigments, food additives and pesticides. The biotechnological tools are important to select, multiply, improve and analyze medicinal plants. In-vitro production of secondary metabolites in plant cell suspension cultures has been reported from various medicinal plants and bioreactors are the key step towards commercial production of secondary metabolites by plant biotechnology. Genetic transformation is a powerful tool for enhancing the productivity of novel secondary metabolites; especially by Agrobacterium tumefacians. Combinatorial biosynthesis is another approach in the generation of novel natural products and for the production of rare and expensive natural products. DNA profiling techniques like DNA microarrays serve as suitable high throughput tools for the simultaneous analysis of multiple genes and analysis of gene expression that becomes necessary for providing clues about regulatory mechanism, biochemical pathways and broader cellular functions.

Keywords: Medicinal plants, biotechnology, combinatorial biosynthesis, DNA microarray, transgenic plants

 

 

Indian Journal of Biotechnology

Vol 8, January 2009, pp 23-32

 

 

Characterization of embryonic stem cells: A special focus on farm animals

 

D Kumar*, T Anand, M K Singh, M S Chauhan and R S Manik

 

Animal Biotechnology Center, National Dairy Research Institute, Karnal 132 001, India

 

Received 12 September 2007; revised 26 May 2008; accepted 17 July 2008

Embryonic stem (ES) cells are derived from the inner cell mass of blastocysts. They grow indefinitely while maintaining the pluripotency in the presence of specific growth factors such as leukemia inhibitory factor. The molecular mechanisms for self-renewal of pluripotent cells and the role of various growth factors involved in self-renewal as well as differentiation are being deciphered. ES cells, in their undifferentiated state, are characterized by a distinct morphology and by the presence of a set of markers classified into intracellular and extracellular types. Expression of specific markers (surface and transcription based) is an important criterion for pluripotent or undifferentiated state of cells. The expression of these markers is found to be exclusive to a particular species. A thorough understanding of the expression of these markers and of factors or conditions for the long-term culture of ES cells, without compromising their pluripotency and a stable genetic make-up is very important for the production and maintenance of ES cells from different species of farm animals. In this review, we present an overview of characterization of embryonic stem cells in farm animals.

Keywords: Embryonic stem cells, characterization, differentiation, karyotyping, epigenetics, farm animals

 

 

Indian Journal of Biotechnology

Vol 8, January 2009, pp 33-39

 

 

Genotype analysis and assessment of antigenic sensitivity for recombinant HCV proteins by indigenous SIBA for detection of Hepatitis C Virus infection: A comparison with 3rd EIA and RT-PCR

 

S P D Ponamgi1, M Chandra1, Y Naresh Kumar1, S Rahamathulla1, Lakshmi Narasu2, C M Habibullah1, and M N Khaja1*

 

1Centre for Liver Research and Diagnostics, Deccan College of Medical Sciences and Allied Hospitals, Kanchanbagh, Hyderabad 500 058, India
2Department of Biotechnology, Jawaharlal Nehru Technology University, Hyderabad 500 072, India

 

Received 26 July 2007; revised 19 May 2008; accepted 5 August 2008

The first serological testing for the detection of anti-HCV antibodies using recombinant antigens was introduced in 1991. Since then many developments have taken place and at present third generation ELISA kits are being used most widely and globally. Detection of anti-HCV does not distinguish past from present infections and in diagnostic virology particularly ELISA’s, a positive HCV test result may be non-specific and therefore has to be crosschecked by another test of different principle for which Immunoblots were initially developed. Patients with liver disease attending the inpatient and outpatient wards of the CLRD (Center for Liver Research and Diagnostics) between Aug 2004 and Feb 2007 were screened for HCV by using 3rd generation ELISA, HCV blot, and RT-PCR. Genotyping was done for all the positive samples. Out of 531 samples tested, 211 samples showed identical results as reactive by ELISA, HCV blot and RT-PCR. Out of the 214 genotype samples, genotype 1a was found to be prevalent by 52.33% (n=112), followed by others. RNA based detection by RT-PCR remains the reliable method of HCV diagnosis, however, where there are no facilities for the PCR to be performed particularly in the small to medium laboratory and diagnostic centers, HCV blot could be done as a supplemental assay.

Keywords: Genotyping, RT-PCR, HCV blot, 3rd generation ELISA

 

 

Indian Journal of Biotechnology

Vol. 8, January 2009, pp 40-45

 

 

Molecular level studies on multiple antibiotic and serum resistance in UTI pathogens

 

S M Dharmadhikari* and S A Peshwe

 

Department of Microbiology, Govt Institute of Science, Caves Road, Aurangabad 431 004, India

 

Received 17 September 2007; revised 5 March 2008; accepted 28 June 2008

A total of six pathogens comprising Escherichia coli (2), Staphylococcus aureus (2) and Proteus sp. (2) were isolated from urine samples of urinary tract infection (UTI) patient. They were examined for antibiotic and serum resistance. E. coli BJ 83, coagulase positive S. aureus and Proteus sp. showed resistance to various antibiotics viz., ampicillin, cefuroxime, streptomycin, etc. as well as resistance to 2% human serum. Amongst them, E. coli BJ 83, which exhibited 62.5% resistance to various antibiotics and serum, was selected for genetic evaluation as well as cotransformation studies. Plasmid curing showed the location of antibiotic resistance markers on R plasmid. The presence of MDR plasmid in E. coli BJ 83 was confirmed by performing RFLP. E. coli BJ 83 plasmid DNA profile showed lmax at 260 nm and Tm of 90oC. Coexpression of ampicillin and serum resistance by recipient strain of E. coli X-239, after plasmid transformation, confirmed that they are found to be linked markers.

Keywords: Antibiotic resistance, serum resistance, virulence, plasmid DNA, RFLP, transformation, E .coli BJ-83

 

 

Indian Journal of Biotechnology

Vol 8, January 2009, pp 46-52

 

 

Structure-function annotation and phylogenetic strategy of nifH domain of a cyanobacterium—Chlorogloeopsis sp.

 

P T V Lakshmi1*, S Uma Maheswari1 and A Annamalai2

 

1Department of Bioinformatics, Bharathiar University, Coimbatore 641 046, India

2Schools of Biotechnology, Institute of Technology and Science, Karunya University, Coimbatore 641 114, India

 

Received 26 March 2007; revised 25 April 2008; accepted 27 June 2008

Exploration of the available protein sequences of the Cyanobacterial genera Chlorogloeopsis of Stigonematales from the NCBI database to annotate structural and functional domains through BLOCKS SEARCHER and SWISS PDB VIEWER (SWISSMODEL) revealed eleven different important domains, of which nitrogen fixing domain showed high frequency of occurrence (18.18%), among the sequences explored. Therefore, it was particularly compared with other protein sequences available in the databases in order to obtain similarity and to evolve a phylogram. Similarity search for the NifH/frxC family sequence using BLAST P showed that it is similar to nitrogenase protein of the genera Mastigocladus, Nostoc, Anabaena and other uncultured cyanobacterium. Phylogenetic tree based on the similarities obtained from BLAST P indicates the evolutionary relationships.

Keywords: BLAST P, Cyanobacteria, Chlorogloeopsis, nifH domain, phylogram

 

 

Indian Journal of Biotechnology

Vol 8, January 2009, pp  53-60

 

 

Down-regulation of an abiotic stress related Nicotiana benthamiana WRKY transcription factor induces physiological abnormalities

 

 

K Archana, N Rama, H M Mamrutha and Karaba N Nataraja*

 

Department of Crop Physiology, University of Agricultural Sciences, GKVK, Bangalore, India

 

Received 24 December 2007 revised 31 July 2008; accepted 3 October 2008

Transcription factors (TFs), the DNA binding proteins, play key role in biotic and abiotic stress responses in plants by regulating the expression of downstream target genes. Many different TFs have been cloned and characterized in model plants and a few of them have been shown to have direct role in abiotic stress tolerance. In the present report, we have cloned a partial cDNA of AtWRKY75 like gene (NbWRKY) from the model plant, Nicotiana benthamiana, and studied its expression under whole plant desiccation (drought) stress. To induce drought stress, soil water status (field capacity, FC) was maintained between 60-65% by controlled irrigation and replacing water transpired twice a day. The extent of drought stress was assessed by monitoring the leaf water status and quantifying photosynthetic pigments. The semi-quantitative RT-PCR revealed constitutive expression of NbWRKY and up-regulation under drought. Down-regulation of NbWRKY by virus induced gene silencing (VIGS) produced chlorosis and senescing phenotype in N. benthamiana. The silenced plants showed reduced photosynthesis, efficiency of open PSII reaction centre, and exhibited the symptoms of photoinhibition. The results indicated the indispensable role of NbWRKY in basic physiological processes.

Keywords: VIGS, transcription factor, WRKY, abiotic stress, gene silencing

 

 

Indian Journal of Biotechnology

Vol 8, January 2009, pp 61-66

 

 

Molecular diversity in genus Nicotiana as revealed by randomly amplified polymorphic DNA

 

K Siva Raju1*, M Sheshumadhav2, C Chandrasekhararao and T G K Murthy2

 

1Division of Crop Chemistry and Soil Science and 2Division of Crop Improvement,
Central Tobacco Research Institute, Rajahmundry 533 105, India

 

Received 7 November 2007; revised 16 April 2008; accepted 25 June 2008

The genus Nicotiana consists of 64 recognized species of which, only 2 species, N. tabacum and N. rustica are cultivated extensively. Wild Nicotiana species are storehouses of genes for several diseases and pests, in addition to genes for several important phytochemicals and quality traits which are not present in cultivated varieties. Randomly amplified polymorphic DNA(RAPD) analysis was used to determine the degree of genetic variation in the genus Nicotiana and to develop species specific markers. 22 species and 2 interspecific hybrids were analyzed by using 18 decamer primers. Greater amount of genetic polymorphism exists among the wild species of genus Nicotiana (99.5%) as evidenced by the high degree of polymorphism in RAPD profiles. The pairwise similarity measures in the species of subgenus Rustica was 0.252 whereas in the subgenus Tabacum it was 0.189 suggesting that there was significant diversity amongst the species of these subgenera. In the species of subgenus Petunioides, the range of pairwise similarity measures was 0.128 to 0.941. The clustering pattern coincided with the traditional classification of Nicotiana species. All the primers generated specific bands in the various species. 36 species specific markers identified in the present study would be utilized in interspecific breeding programmes.

Keywords: Nicotiana, genetic diversity, RAPD

 

 

Indian Journal of Biotechnology

Vol. 8, January  2009, pp 67-71

 

 

Use of RAPD markers for assessment of genetic diversity in sugarcane cultivars

 

P G Kawar, R M Devarumath* and Y Nerkar

 

Molecular Biology and Genetic Engineering Laboratory, Vasantdada Sugar Institute, Pune 412 307, India

 

Received 27 April 2007; revised 20 August 2008; accepted 13 October 2008

Random amplified polymorphic DNA (RAPD) analysis was carried out in 17 cultivars of sugarcane. Selected 40 primers generated 325 bands, 134 of which were found to be polymorphic. The number of amplification products ranged from 3 to 15 for different primers. The genetic similarity among sugarcane cultivars ranged from 0.77 to 0.99. The average genetic similarity was 0.87. UPGMA cluster analysis placed these cultivars in to different groups. The parentage of the varieties did not contribute significantly to the grouping pattern. Varieties belonging to the common female parent were under different groups, while varieties from different parentage were under same group. Among the varieties Co SNK 3754 was found to be totally distinct and divergent from rest of the varieties. The study reveals the limited genetic base of the current Indian commercial varieties and the need to diversify the genetic base by using new sources from the germplasm.

Keywords: Genetic diversity, RAPD markers, sugarcane

 

 

Indian Journal of Biotechnology

Vol 8, January 2009, pp 72-77

 

 

Enhanced insect resistance to bollworm (Helicoverpa armigera) in cotton containing a synthetic cry1Ab gene

 

Farzane Yazdanpanah1, M Tohidfar1*, M Esna Ashari2, B Ghareyazi1, M Karimi Jashni3 and M Mosavi 1

 

1Agricultural Biotechnology Research Institute of Iran (ABRII), P O Box 31535-1897, Mahdasht Road, Karaj, Iran

2University of Hamedan, Department of Biotechnology and Plant Breeding, Hamedan, Iran

3Iranian Research Institute of Plant Protection, P O Box 19395-1454, Tabnak St., Tehran, Iran

 

Received 22 June 2007; revised 21 May 2008; accepted 2 August 2008

In order to investigate stability of the cry1Ab gene and resistance in the T1 generation of transgenic cotton (Gossypium hirsutum var. Coker 100), transgenic seeds were analyzed by PCR, Southern and Western blotting and bioassayed. Seed samples from transgenic lines (T0) along with seeds from non-transgenic lines s as control were used. PCR analysis showed the presence of cry1Ab gene in 63 out of the 150 T1 plants. Integration of the cry1Ab gene into the genome of transgenic plants was confirmed by Southern blotting. Western immunoblot analysis of transgenic plants revealed the presence of a band with MW of 67 kDa using anti- Cry1Ab-polyclonal antiserum. Bioassay tests indicated that the Cry1Ab protein was active and that the larvae died in the first days. Significant differences were observed in the mortality rate, growth rate and extent of damage on the leaves between the transgenic and non-transgenic plants. The average mortality of the larvae in line 17 was 100%, in comparison with the control. These results indicated the relative advantage of line 17, which carries one copy of the cry1Ab gene in comparison with the other lines.

Keywords: Insect resistant cotton, transgenic plant, cry1Ab gene, second generation, Helicoverpa armigera.

 

 

Indian Journal of Biotechnology

Vol 8, January 2009, pp 78-84

 

 

Agrobacterium–mediated transformation of chickpea using shoot meristem

 

Rekha Singh, N P Singh*, Subhojit Datta, Indu Singh Yadav and A P Singh

 

Biotechnology Unit, Indian Institute of Pulses Research, Kanpur 208 024, India

 

Received 7 September 2006; revised 23 July 2008; accepted 24 September 2008

Agrobacterium-mediated gene transfer to pre-organized meristematic tissue combined with axillary regeneration was standardized for transformation and regeneration of chickpea, which otherwise was difficult to achieve from other explants. Different Agrobacterium strains harbouring binary vectors pCGP1258, containing the GUS as a reporter and bar [gene for resistance to phosphinothricin (PPT)—the active ingredient of the herbicide Basta] as the selectable marker, were used for the transformation experiments. After co-cultivation, the shoot apex explants were transferred onto a PPT-free regeneration medium and their tops (2 mm) were thoroughly wetted with PPT solution (2 mg/mL). The multiple axillary shoots developing from the shoot apices were excised and placed onto a medium containing 10 mg/L PPT. The surviving shoots were subcultured every 2nd wk onto fresh medium containing 20 mg/L PPT. After each subculture, the number of surviving shoots decreased until it stabilized. Some of the chimeric shoots surviving the PPT selection eventually developed new healthier axillary shoots, which could be rooted or grafted on in vitro grown seedling. This whole process took 6-9 months. Average transformation frequency was found between 1.29-3.33%. Transmission of the transgenes into progeny was also studied following the inheritance of uid A gene in T1 and T2 progenies. The overall segregation ratio among progenies of plants derived from T0 plants appeared to be close to 3:1 Mendelian ratio, indicating integration of the transgene at single locus.

Keywords: Agrobacterium tumefaciens, chickpea, Cicer arietinum, genetic transformation, β-glucuronidase

 

 

Indian Journal of Biotechnology

Vol. 8, January 2009, pp 85-90

 

 

Genetic variation and differentiation in the Stinging catfish, Heteropneustes fossilis (Bloch), populations assessed by heterologous microsatellite DNA markers

 

Shamima Nasren, Mohammad Nazrul Islam1, Mohd Golam Quader Khan, Md Shahidul Islam1 and Md Samsul Alam*

 

Department of Fisheries Biology and Genetics and  1Department of Biotechnology, Bangladesh Agricultural University,
Mymensingh 2202, Bangladesh

 

Received 21 February 2008 ; revised 11 August 2008; accepted 15 October 2008

Microsatellite DNA markers have been increasingly used in genetic diversity studies. The present study reports on the characterization of genetic variation and differentiation in four different natural populations of the stinging catfish, Heteropneustes fossilis (Bloch), in Bangladesh, viz., Mymensingh, Netrakona, Narsingdi and Rangpur, using cross-species microsatelllite DNA markers developed from the walking catfish, Clarias batrachus. Eighteen polymorphic alleles were found in the 128 diploid individuals (32 from each population), with nine alleles at each of the two loci analyzed. The Netrakona and Rangpur population deviated from the Hardy-Weinberg proportion at one locus. The population differentiation (FST) value between the Narsingdi and Netrakona population was found to be insignificant, while the values between all the other population pairs were found to be significant. The genetic distance values ranged between 0.165 and 0.626. The UPGMA dendrogram based on genetic distance resulted in two clusters: the Mymensingh population alone was in one cluster and the three other populations in the second cluster. This study revealed a fairly high level of genetic variation in the microsatellite loci within and between the four populations, and identified existence of distinct population groups of H. fossilis.

Keywords: Genetic variation, Heteropneustes fossilis, microsatellite, polymorphism

 

 

Indian Journal of Biotechnology

Vol 8, January 2009, pp 91-96

 

 

Probable mechanism(s) of antifungal activity of HA-2-91—A tetraene antibiotic

 

S R Naik* and T E Gupteψ

 

Research and Development Centre, Hindustan Antibiotics Ltd, Pimpri, Pune 411 018, India

 

Received 11 October 2006; revised 12 May 2008; accepted 4 August 2008

HA-2-91 (a tetraene antibiotic) is produced by Streptomyces arenae var. ukrainiana and shown to elicit antifungal activity (in vitro) against yeasts and filamentous fungi including plant pathogens and clinical isolates. Experimental studies were carried out by using both biological as well as chemical methods to understand the probable mechanism(s) of antifungal action of HA-2-91. Experimental findings suggest that HA-2-91 binds to both ergosterol and cholesterol present on the fungal and mammalian cell membranes. However, results showed preferential binding of HA-2-91 towards ergosterol than cholesterol. Further, the interactions (bindings of HA-2-91 with membrane sterols) seem to be reversible process. With such preferential degree of binding of HA-2-91, the effect will be greater on fungal cell membrane than mammalian cell membrane. Hence, adverse reactions like haemolysis may not occur.

Keywords: Antifungal action, affinity for sterols, reversible binding process, tetraene.

 

 

Indian Journal of Biotechnology

Vol 8, January 2009, pp 97-109

 

 

Characterization of functional activity in composted casing amendments used in cultivation of Agaricus bisporus (Lange) Imbach

 

Devendra K Choudhary*, Pavan K Agarwal1, and Bhavdish N Johri1

 

Department of Microbiology, G B Pant University of Agriculture and Technology, Pantnagar 263 145, India
1Department of Biotechnology, Barkatullah University, Bhopal 462 026, India

 

Received 23 August 2007; revised 20 August 2008; accepted 16 October 2008

In cultivation of button mushroom [Agaricus bisporus (Lange) Imbach], casing layer that is nutritionally deficient to compost is believed to trigger the fruit body formation and this is conducted by the bacterial community residing in casing layer. The change in nutritional status of the casing is highly correlated with microbial flora. Therefore, an attempt was made to characterize the bacterial flora in casing layer, i.e., Farm Yard Manure and Spent Mushroom Substrate (FYM+SMS, 3:1) and Farm Yard Manure and Vermi Compost (FYM+VC, 3:1), employing phenetic approaches. Morphotypically different and functionally characterized bacterial isolates were identified by partial 16S rDNA gene fragment sequencing. Available data showed a significant variety of organisms that included Acinetobacter and Pseudomonas of the γ-proteobacteria, that were most frequently encountered genera. Amongst Gram-positive bacteria, Bacillus was the most highly represented genus that was derived from the agaric fruit bodies. In addition, FYM+SMS was found to be a high yielding casing mixture, which took minimum case run period together with superior fruit body quality as compared to FYM+VC.

Keywords: Acinetobacter, Bacillus, casing, endotrophs, phenetic approach, Pseudomonas

 

 

Indian Journal of Biotechnology

Vol 8, January 2009, pp 110-114

 

 

Characterization of thermoalkalophilic xylanase isolated from Enterobacter isolates

 

Anjana Sharma*, Rajesh Pujari and Pratibha Patel

 

Bacteriology Laboratory, Department of P G Studies and Research in Biological Sciences
R D University, Jabalpur 482 001 India

 

Received 15 January 2008, revised 20 August 2008, accepted 23 October 2008

Thermoalkalophilic xylanase was isolated from Enterobacter spp. with significant activity. Maximum enzyme activity was observed in isolate BGCC#259 (E. cloacae), i.e., 0.056 IU/mL at 24 h, while the highest biomass was observed at 36 h of incubation. Hydrolysis study revealed complete degradation of xylan to xylose after 12 h of incubation. The molecular mass of the xylanase was ~43 kDa. The enzyme was characterized at varied range of pH and temperature, and it was found that all five Enterobacter isolates had a pH and temperature optima of 8.0 and 80oC, respectively. At 80oC, xylanase from isolate BGCC#254 (E. cloacae) retained 100% activity, while BGCC#259 retained more than 90% activity after 24 h.

Keywords: Characterization, Enterobacter, enzyme activity, thermoalkalophilic, xylanase

 

 

Indian Journal of Biotechnology

Vol 8, January 2009, pp 115-120

 

 

Micropropagation of Wrightia tomentosa : Effect of gelling agents, carbon
source and vessel type

 

Preeti Joshi, Rohini Trivedi and S D Purohit*

 

Plant Biotechnology Laboratory, Department of Botany, Mohanlal Sukhadia University, Udaipur 313 001, India

 

Received 23 August 2007; revised 16 April 2008; accepted 22 June 2008

The role of gelling agents, carbon source, type of culture vessel and liquid culture system during rooting phase in Wrightia tomentosa was investigated in an effort to reduce the cost of micropropagation. Studies have revealed significant improvement in shoot multiplication on a medium containing crude agar and sugarcubes. Promotory role of vented vessel in culture growth and liquid culture system during rooting phase was established. The results showed strong potential for reduction in cost of plantlets in vitro.

Keywords: Micropropagation, vented vessels, hyperhydration, Wrightia tomentosa

 

 

Indian Journal of Biotechnology

Vol 8, January 2009, pp 121-126

 

 

Regeneration competence of Tainia latifolia (Lindl.) Benth ex Hook pseudobulb segments: An in vitro study

 

Sungkumlong and Chitta Ranjan Deb*

 

Department of Botany, Nagaland University, Headquarters: Lumami, Mokokchung 798 601 (University Branch), Nagaland, India

 

Received 28 December 2007; revised 25 August 2008; accepted 24 October 2008

Pseudobulb segments (~5 mm size) obtained from in vitro raised plantlets as well as greenhouse grown Tainia latifolia (Lindl.) Benth ex Hook were inoculated on different basal media, viz., MS, Mitra et al and Knudson ‘C’ with different levels of adjuncts. The explants from in vitro source out performed the in vivo sourced pseudobulb segments as for as initial morphogenetic response is concerned. Of the three basal media studied, MS medium in conjunction with 3% sucrose, 100 mg L-1 casein-hydrolysate (CH), 100 mg L-1 citric acid (as antioxidant) and 0.1% activated charcoal (AC) performed better. Within 7 wk of culture, the explants formed as many as 12 shoot buds on MS medium containing 3 µM NAA +  6 µM BA, where about 78% explants responded positively. While 3-4 shoot buds developed after 10 wk of culture from the in vivo sourced explants on MS medium containing 3 µM NAA + 12 µM BA and about 72% explants exhibited positive response. Incorporation of AC in the media proved to be promotory by reducing the browning of the medium and early initiation of the culture. The shoot buds from the initiation medium converted into rooted plantlets within 2-3 passages on MS medium containing 3% sucrose, 0.1% AC, 100 mg L-1 citric acid and 3 µM NAA + 9 µM BA. The well rooted plantlets (~10 cm long with 2 or 3 roots) were hardened for 10-12 wk in culture vials containing 1/10th MS salt solution containing 2% sucrose, pieces of charcoal, coconut husk and chopped litter (1:1:1). The hardened plants were transferred to community potting mix and maintained in the poly-house for 7-8 wk before transferring to the wild. The transplants registered about 70% survival after two months of transfer.

Keywords: Activated charcoal, plant regeneration, pseudobulb culture

 

 

Indian Journal of Biotechnology

Vol 8, January 2009, pp 127-131

 

 

Regeneration of Biophytum sensitivum (Linn.) DC. through organogenesis and somatic embryogenesis

 

M B Shivanna*, M M Vasanthakumari and M C Mangala

 

Department of P G Studies in Applied Botany, Kuvempu University, Shankaraghatta 577 451 India

 

Received 5 October 2007; revised 25 April 2008; accepted 4 July 2008

A protocol was established to regenerate Biophytum sensitivum (L.) DC. through indirect and direct organogenesis and somatic embryogenesis. In the present study, MS medium supplemented with 2,4-D or NAA in combination with BAP induced callusing in stem, inflorescence tip and flower bud explants. MS amended with BAP caused callusing in stem and flower bud explants but failed to cause callusing in inflorescence tip explants. Calli obtained on 2, 4-D or BAP amended media produced multiple shoots upon subculturing on BAP (1.0 mg L-1). When a combination of NAA (0.5 mg L-1) + BAP (2.0 mg L-1) was used calli obtained from stem, inflorescence tip and flower bud explants produced multiple shoots. In case of direct organogenesis, only inflorescence tip explants produced multiple shoots on MS medium supplemented with BAP. BAP at 3.0 mg L-1 induced maximum (50%) response. Regenerated shoots rooted on half strength MS medium supplemented with NAA 0.5 mg l-1. MS medium supplemented with 2, 4-D (2.5 mg l-1) induced embryogenic response in calli from inflorescence explants. Embryogenic calli produced embryoids upon transfer to MS medium supplemented with NAA (1.0 mg L-1) + BAP (3.0 mg L-1). Embryoids were germinated on half strength MS medium. Eighty per cent of the rooted plantlets and 90% of somatic embryo derived plantlets survived on soil medium.

Keywords: Biophytum sensitivum, in vitro regeneration, organogenesis, somatic embryogenesis

 

 

Indian Journal of Biotechnology

Vol 8, January 2009, pp 132-135

 

 

Direct organogenesis and somatic embryogenesis in Beloperone plumbaginifolia (Jacq.) Nees

 

M C Shameer, V P Saeeda, P V Madhusoodanan and Sailas Benjamin*

 

Biotechnology Division, Department of Botany, University of Calicut, Kerala 673 635, India

 

Received 27 December 2007; revised 18 August 2008; accepted 22 October 2008

Beloperone plumbaginifolia (Jacq.) Nees is a small branched medicinal shrub. In vitro studies were conducted, employing explants from node, internode, petiole, shoot bud and leaf lamina. Murashige and Skoog (MS) medium fortified with 6.66 μM BA enabled the proliferation of axillary and apical buds. MS medium supplemented with 5.37 mM NAA was better for callogenesis from nodal and internodal explants, while a combination of IBA (2.46 mM) and 2,4-D (4.52 mM) was good for leaf lamina explant. MS medium with 5.37 mM NAA and 2.22 mM BA was found superior for shoot induction from nodal explants. Half-strength MS medium with 5.37 mM NAA induced adventitious roots and 85% plantlets survived when transferred in the field conditions.

Keywords: Beloperone plumbaginifolia, callogenesis, organogenesis, somatic embryogenesis.