Volume 1, Number 1,    June 2005

Table of contents          Abstract

Abstract


Biopreparations for plant protection in Siberia : application and enhancement of activity
 Margarita Shternshis *

Department for Biological Control, Novosibirsk State Agrarian University 630039, Novosibirsk , Russia

Shternshis, M. (2005). Biopreparations for plant protection in Siberia : application and enhancement of activity. Journal of Agricultural Technology 1 (1) : 1-18.
This review is devoted to the most widespread biological preparations used for ecologically safe control of insect pests and plant diseases in Siberia . Special attention is given to the application of biopreparations on vegetable and soft fruit crops. The necessity of enhancement of control agent activity and relevant formulations is discussed. In most cases, data obtained by Siberian authors have been presented. Possible ways to enhance efficiency of biopreparations using the mixtures based on biocontrol agents of different origin or on the agents with nontoxic enhancers are shown. The role of host plant in the efficacy of biopreparations is emphasized, especially for future research.

Key words: biocontrol, biopreparation, enhancer, insect pest, plant disease.

* e-mail: shternshis@pochta.ru , shternshis@mail.ru

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Application of mycofungicide to control late blight of potato

Kasem Soytong 1 * and Kanchalika Ratanacherdchai 1

1 Department of Plant Pest Management Technology, Faculty of Agricultural Technology, King Mongkut’s Institute of Technology Ladkrabang (KMITL), Bangkok 10520,Thailand

Soytong, K. and Ratanacherdchai, K. (2005). Application of mycofungicide to control late blight of potato. Journal of Agricultural Technology 1 (1) : 19-32.
Late blight of potato in northern Thailand is caused by Phytophthora infestans and leads to economic damage in large areas of potato planted. The pathogen infects all stages of plant growth which show symptoms of late blight, stem rot and tuber rot and can result in 100% yield loss even with the use of chemical fungicides. Laboratory based bi-culture antagonistic tests showed that Chaetomium-mycofungicide inhibited pathogen growth by more than 53% over 10 days. In field trials, we used a Chaetomium-mycofungicide for disease control in combination with biological fertilizers comprising 12 effective strains of cellulose degrading fungi and specific fungi for plant growth stimulants. The mixture significantly reduced disease indices in infected fields when compared with the non-treated control and the difference between Chaetomium-mycofungicide and chemical pesticide treated fields was not significant. Three Chaetomium-mycofungicide/biological fertilizer treatments were tested and compared with no treatment and a chemical pesticide treatment. Bio-technique 1 reduced late blight incidence by 34% and pathogen colonization by 56.5% leading to an increase in yield of 49%. Bio-technique 2 reduced late blight incidence by 15.5% and reduction colonization by 29% leading to an increased yield of 49%. Bio-technique 3 reduced late blight incidence by 19% and pathogen colonization by 29% leading to an increased yield of 45.5%. Chemical pesticide treatment reduced late blight incidence by 34.5% and pathogen colonization of 51.5% leading to an increased yield of 52.5%. Non-treatment gave the lowest yield due to high late blight incidence. It is recommended that the use of Bio-IPM techniques could solve the problem of disease epidemics in the infested areas planted to potato in the cool tropics.

Key words: biological fertilizers, Chaetomium, late blight of potato, mycofungicide

* Corresponding author: Kasem Soytong: e-mail: kskasem@kmitl.ac.th

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Application of antagonistic fungi to control anthracnose disease of grape

K. Soytong 1 *, W. Srinon 2 , K. Rattanacherdchai 1, S. Kanokmedhakul 3 and K. Kanokmedhakul 3

1 Department of Plant Pest Management, Faculty of Agricultural Technology, King Mongkut’s Institute of Technology Ladkrabang, Bangkok 10520, Thailand
2 Department of Plant Science, Maejo University, Phrae 54140, Thailand
3 Department of Chemistry, Faculty of Science, Khon Kaen University, Khon Kaen 40002, Thailand

Soytong, K., Srinon, W., Rattanacherdchai, K., Kanokmedhakul, S. and Kanokmedhakul, K. (2005). Application of antagonistic fungi to control anthracnose disease of grape. Journal of Agricultural Technology 1 (1) : 33-42.
Anthracnose of the grape varieties Bigblack, Nanpha, Blackopal, Loose perlette and White malaca are caused by Colletotrichum gloeosporioides. All isolates obtained from grape anthracnose were shown to be pathogenic; isolate WMF01 was the most virulent on all tested varieties of grape. Assays using crude extracts from Chaetomium cupreum CC, C. globosum CG, Trichoderma harzianum PC01, T. hamatum PC02, Penicillium chrysogenum KMITL44 and antibiotic substances Rotiorinol, Chaetoglobosin-C and Trichotoxin A50 were carried out to test bioactivity. All extracts and compounds inhibited the growth of C. gloeosporioides strain WMF01, with average ED50 values between 1 to 50 ppm. Applications of bioproducts of Chaetomium, Penicillium and Trichoderma, and a mixture of those bioproducts in a powder formulation and a chemical control were conducted in the field to control anthracnose disease of 5-varieties of grape. All bioproducts significantly reduced the disease incidence on leaves, twigs and fruits of grape in all varieties as compared to the chemical control.

Key words: antagonistic fungi, Chaetoglobosin-C, Chaetomium, Penicillium, Rotiorinol, Trichoderma, Trichotoxin A50

* Corresponding author: Kasem Soytong: e-mail: kskasem@kmitl.ac.th

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Anti-feedant and growth inhibitory effects of seed extracts of custard apple, Annona squamosa against Khapra beetle, Trogoderma granarium

N.S. Rao *, K. Sharma and R.K. Sharma

Division of Entomology, Indian Agricultural Research Institute, New Delhi – 110012, India

Rao, N.S., Sharma, K. and Sharma, R.K. (2005). Anti-feedant and growth inhibitory effects of seed extracts of custard apple, Annona squamosa against Khapra Beetle, Trogoderma granarium. Journal of Agricultural Technology 1 (1) : 43-54.
The anti-feedant and growth inhibitory effects of seed extracts of custard apple in hexane, ethyl acetate and methanol were tested against neonate and 7 days old larvae of the Khapra beetle, Trogoderma granarium, using a feeding bioassay. The LC50 values for hexane, ethyl acetate and methanol extracts were 1195.41, 305.36 and 1446.32 ppm for neonate larvae; 5805, 1300 and 5815 ppm for 7 days old larvae, respectively. Drastic reduction in larval weight was observed in all extracts as compared with the control (acetone). The weight of five larvae in the control (acetone) was 6.03 mg after 15 days of release of neonate larvae. In case of Annona squamosa seed hexane extract the larval weight near LC50 (1250 ppm) was 0.65 mg, whereas in ethyl acetate (250 ppm) and methanol extract (1500 ppm) it was 0.91 and 0.90 mg, respectively. Ethyl acetate extract at 1250 ppm showed maximum anti-feedant activity of 44.50 and 59.50% after 10 and 15 days of release, respectively, whereas 35.68 and 53.45 % for hexane extract and 26.07 and 38.65% for methanol extract was observed at same concentration after 10 and 15 days of release, respectively. Annona squamosa seed ethyl acetate extract produced 55.73% anti-feedant activity for 7 days old larvae while hexane and methanol extracts produced 50 and 25.04%, respectively. The present study conclusively showed that A. squamosa seed ethyl acetate extract was superior to hexane and methanol extracts and first such study against Khapra beetle.

Key words:Annona Squamosa, anti-feedant activity, seed extracts

* Corresponding author: N.S. Rao; e-mail:getsambuat@yahoo.co.in

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Enzyme production by endophytes of Brucea javanica

Y.W. Choi, I.J. Hodgkiss and K.D. Hyde *

Centre for Research in Fungal Diversity, Department of Ecology & Diversity, The University of Hong Kong, Pokfulam Road, Hong Kong SAR, PR China

Choi, Y.W., Hodgkiss, I.J and Hyde, K.D. (2005). E nzyme production by endophytes of Brucea javanica. Journal of Agricultural Technology 1 (1) : 55-66.
Twenty-one endophytic isolates from Brucea javanica were tested for their ability to produce extracellular cellulase and extracellular and intracellular amylase, ligninase, pectinase and xylanase. The same fungi were tested for their ability to cause weight loss in wood blocks. All fungi produced amylase and cellulase, while only one sterile mycelium produced ligninase and no isolates produced pectinase. The enzyme tests indicate that most endophytes are degraders of the simpler sugars and cellulose available in recently dead leaves and possibly wood. Only one slow growing species of sterile mycelium however, appeared to be capable of degrading lignin that would be available in dead wood. No fungi appeared to be latent pathogens. A discussion of enzyme production in relation to possible roles of endophytes is provided.

Key words: endophytes, enzyme production, latent pathogens, ligninases.

* Corresponding author: K.D.Hyde; e-mail: kdhyde@hkucc.hku.hk

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Antimicrobial and enzyme activity of mangrove endophytic fungi of southwest coast of India

G.L. Maria 1, K.R. Sridhar 2 * and N.S. Raviraja 2

1 Department of Botany, St. Agnes College , Mangalore 575 002, Karnataka , India
2 Microbiology and Biotechnology, Department of Biosciences, Mangalore University , Mangalagangotri, Mangalore 574 199, Karnataka , India

Maria, G.L., Sridhar, K.R. and Raviraja , N.S. (2005). Antimicrobial and enzyme activity of mangrove endophytic fungi of southwest coast of India. Journal of Agricultural Technology 1 (1) : 67-80.
The antimicrobial potential of 14 endophytic fungi isolated from Acanthus ilicifolius and Acrostichum aureum towards selected bacteria (Bacillus subtilis, Enterococcus sp., Klebsiella pneumoniae, Pseudomonas aerugionsa, Salmonella typhi and Staphylococcus aureus) and fungi (Candida albicans and Trichophyton metagrophytes) was tested using ethyl acetate extract of fungi cultivated under solid-state fermentation (SSF). Aspergillus spp. showed promising antagonistic features. All test bacteria were inhibited by sterile isolate MSI 1. Cumulospora marina and Pestalotiopsis sp. showed considerable inhibition on Gram-positive and Gram-negative bacteria. Crude ethyl extracts derived from submerged fermentation (SmF) of four endophytes showing positive results were further evaluated. Aspergillus sp. 3 and Pestalotiopsis sp. inhibited many bacteria and Candida albicans, while inhibition by Aspergillus sp. 2 and MSI 1 was confined to bacteria. Crude ethyl acetate extracts purified by TLC of four endophytes grown under SmF showed many fluorescent fractions. Two fractions of Aspergillus sp. 3 showed high antimicrobial activity. One of the fractions of Pestalotiopsis sp. showed considerable inhibition of Bacillus subtilis, Staphylococcus aureus and Candida albicans. Seven endophytic fungi were assessed for the production of extracellular enzymes (amylase, cellulase, chitinase, laccase, lipase, protease and tyrosinase) by culture plate method. Cellulase and lipase activity was present in all fungi, while amylase and protease in a few. No fungi exhibited chitinase, laccase and tyrosinase activity. Enzyme production of Pestalotiopsis sp. (cellulase by SmF; xylanase, pectinase and protease by SSF) at pH 7 and pH 9 during 3-15 days of fermentation was assessed. The Cellulase activity was highest at pH 7 on 6th day, while xylanase activity was highest on 9th day at pH 9. Highest activity of pectinase was seen on 6th (pH 7) and 9th days (pH 9). Protease activity was highest on 6th day at pH 7 and 9.

Key words: antimicrobial activity, endophytic fungi, enzymes, mangroves.

* Corresponding author: e-mail: sirikr@yahoo.com

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Evaluation of anamorphic state, wood decay and production of lignin-modifying enzymes for diatrypaceous fungi from Argentina

 M.B. Pildain, M.V. Novas and C.C. Carmarán *

Deptmento de Biodiversidad y Biología Experimental, Lab. Micología-PHRIDEB, CONICET, Fac. de Cs. Exactas y Naturales, Universidad de Buenos Aires, Capital Federal EHA1428, Argentina

Pildain, M.B., Novas, M.V. and Carmarán, C.C. (2005). Evaluation of anamorphic state, wood decay and production of lignin-modifying enzymes for diatrypaceous fungi from Argentina. Journal of Agricultural Technology 1 (1) : 81-96.
Morphological descriptions, wood decay and anatomical criteria, production of ligninolytic and cellulolytic enzymes and melanin production of diatrypaceous species (Eutypella andicola, E. comosa, E. leprosa and E. scoparia) from Argentina were investigated. The anamorphic state of Eutypella andicola is described for the first time. The morphological characteristic of the anamorphs were similar among all strains. Most isolates were capable of causing mass loss in Populus deltoides wood blocks during a 12-week period. Differences among strains were significant. The anatomical observations demonstrated that in a first state of wood colonisation this fungi produced soft rot decay and in later colonisation state the decay type is similar to white rot. Extracellular production of ligninolytic and cellulolytic enzymes were studied by mycelium growing on solid medium supplemented with different dyes (Malachite green, Azure B, Poly R-478, Congo red, tannic and galic acid, and guayacol). Seven of the eight strains analysed decolourised all the dyes except for the Malachite green. Only one strain of E. scoparia was able to decolourise Malachite green. Through simple assays we established the production of “extracellular melanins” and its association to growth areas of the fungus and discoloration zones.

Key words:Diatrypaceae, Eutypella, ligninolytic enzymes, white rot fungi.

* Corresponding author: C.C. Carmarán ; e-mail: carmaran@bg.fcen.uba.ar

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Determination of ploidy of a dimorphic zygomycete Benjaminiella poitrasii and the occurrence of meiotic division during zygospore germination  

V. Ghormade 1 *, P. Shastry 2, J. Chiplunkar 2 and M.V. Deshpande 1*

1 Biochemical Sciences Division, National Chemical Laboratory, Dr Homi Bhabha Road , Pune-411008, India
2 National Center for Cell Sciences, University of Pune campus, Ganeshkhind, Pune –411007, India

Ghormade, V., Shastry, P., Chiplunkar, J. and Deshpande, M.V. (2005). Det ermination of ploidy of a dimorphic zygomycete Benjaminiella poitrasii and the occurrence of meiotic division during zygospore germination. Journal of Agricultural Technology 1 (1) : 97-112.
Benjaminiella poitrasii is a zygomycetous, dimorphic fungus which exists in yeast or hyphal forms during the vegetative phase and produces asexual sporangiospores and zygopores during the reproductive stage. The zygospores germinate either by germ-sporangiophore formation or hyphal formation. However, the budding type germination of zygospores of B . poitrasii was observed in response to high glucose, 37 ° C and pH 4.0; conditions favouring the yeast-form. The yeast cells from the budding zygospore were analysed to ascertain time and occurrence of meiotic division and to understand the change in the ploidy levels in its life-cycle. The ploidy and nuclear behaviour of this fungus were studied at different stages in the life cycle using the vegetative yeast cells, the asexual sporangiospores, and the yeast cells from the budding zygospore. The uninucleate sporangiospores and the multinucleate yeast cells showed similar DNA contents/nucleus as estimated by spectrophotometric DNA content estimation, survival in the presence of ultraviolet radiation and flow cytometry. The sporangiospores and yeast cells were in the haploid state. DAPI staining of the zygospore showed that few nuclei fused in the zygospore and underwent the meiotic division. The haploid nature of the yeast cells from the budding zygospore indicated that meiosis had occurred before the initiation of germination. The understanding of spore germination in B. poitrasii and its use as a model to screen anti-fungal agents is also discussed.

Key words: B. poitrasii ,DAPI staining, ploidy, DNA contents, flow cytometry

* Corresponding Author: M.V. Deshpande; e-mail: mvdesh@dalton.ncl.res.in

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Production potential of water-soluble Monascus red pigment by a newly isolated Penicillium sp.

Y. Jiang 1, H.B. Li 2 , F. Chen 2 and K.D. Hyde 3 *

1 Department of Biology, Hong Kong Baptist University , Kowloon Tong, Hong Kong SAR, PR China
2 Department of Botany, The University of Hong Kong , Pokfulam Road , Hong Kong SAR, PR China
3 Center for Research in Fungal Diversity, Department of Ecology & Biodiversity, The University of Hong Kong , Pokfulam Road , Hong Kong SAR, PR China

Jiang, Y., Li, H.B., Chen, F. and Hyde, K.D. (2005). Production potential of water-soluble Monascus red pigment by a newly isolated Penicillium sp. Journal of Agricultural Technology 1 (1) : 113-126.
A new isolate from medicinal plant endophytes was identified as Penicillium sp. (HKUCC 8070). A water-soluble red pigment was produced by this strain in potato-dextrose broth, malt-extract broth and a chemically defined medium containing glutamate as a nitrogen source. The red pigment produced was identified as heat-stable, polyketide Monascus red pigment. The highest yield of the red pigment was 1107 mg l-1 obtained from the culture of Penicillium sp. (HKUCC 8070) grown on the malt-extract medium.

Key words: biotechnology, endophytes,fungal pigments,

* Corresponding author K.D. Hyde; e-mail:kdhyde@hkucc.hku.hk

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Application of pectic zymogram in the identification and genetic variation of Fusarium species

G.R. Balali * and M. Iranpoor

Department of Biology, University of Isfahan , Isfahan , IRAN

Balali, G.R. and Iranpoor, M. (2005). A pplication of pectic zymogram in the identification and genetic variation of Fusarium species. Journal of Agricultural Technology 1 (1) : 127-143.
The genus Fusarium is a species rich genus. Different media are required to study the morphological characters and classify different species, and this is a time consuming technique. Molecular and biochemical techniques have been used for identifying fungi over the last two decades, including pectic zymograms which have been used to characterize different fungi. In this study pectic zymograms were used to identify species and form species of Fusarium isolates. Fusarium isolates (318) were obtained from different areas and hosts in Isfahan Province , Iran . Isolates were identified to species based on morphological characters. A pectic enzyme solution was prepared for each isolate using liquid media containing citrus pectin as the sole carbon source. Electrophoresis was performed using acrylamide gel containing 0.2% citrus pectin as the enzyme substrate. The gels were incubated in 0.1M malic acid before staining overnight in 0.02% ruthenium red, to visualize enzyme electrophoretic patterns. Several zymogram phenotypes were obtained for polygalacturonase and pectin esterase. In total, 12 zymogram patterns were determined for the 318 isolates tested. The results showed that there is considerable intraspecific variation in Fusarium species. There were 3, 5 and 2 zymogram electrophoretic patterns for Fusariumoxysporum, F. solani and F. culmorum respectively. However, there were only one zymogram pattern for F. subglutinans and also one for F. equiseti. Although the intraspecific variation based on pectic zymograms was not correlated to the form species of Fusarium, species of Fusarium could be distinguished using this technique as there was no common zymogram pattern among species.

Key words: electrophoresis,pectic enzymes, taxonomy, zymograms

*Corresponding author: G.R. Balali; e-mail: rbalali@yahoo.com

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Identification of fungal phyto-pathogens by Fourier-transform infrared (FTIR) microscopy

V. Erukhimovitch1 * , L. Tsror2, M. Hazanovsky2, M. Talyshinsky1, I. Mukmanov1, Y. Souprun1 and M. Huleihel1

1The Institute for Applied Biosciences, Ben-Gurion University of the Negev, P.O. Box 653, Beer-Sheva 84105
2Department of Plant Pathology, The Institute of Plant Protection, Agricultural Research Organization, Gilat Experiment Station, M.P. Negev, 85250, Israel

Erukhimovitch, V., Tsror, L., Hazanovsky, M., Talyshinsky, M., Mukmanov, I., Souprun, Y.and Huleihel, M. (2005). Identification of fungal phyto-pathogens by Fourier-transform infrared (FTIR) microscopy. Journal of Agricultural Technology 1 (1) : 145-152.
Fungi are considered as serious pathogens to many plants and can cause a severe economic damage. Early detection and identification of these pathogens is very important and might be critical for their control. The available methods for identification of fungi are time consuming and not always very specific. Fourier-transform infrared (FTIR) microscopy is proved to be a reliable and sensitive method for detection of molecular changes in cells. In the present study we used FTIR microscopy as a sensitive and effective assay for the detection and discrimination between different genera of fungi. Our results showed significant spectral differences between the various examined fungal genera.

Keywords: FTIR microscopy, fungal detection, fungi, spectral characteristics

* Corresponding author: V. Erukhimovitch; e-mail: evitaly@bgumail.bgu.ac.il

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Viability in spawn stocks of the white button mushroom, Agaricus bisporus, after freezing in liquid nitrogen without a cryoprotectant

G. Mata 1 *and A.E. Rodríguez Estrada 1

1 Instituto de Ecología, Unidad de Micología , Apartado Postal 63, Xalapa 91000, Veracruz, Mexico

Mata, G. and Rodríguez Estrada A.E. (2005). Viability in spawn stocks of the white button mushroom, Agaricus bisporus, after fre ezing in liquid nitrogen without a cryoprotectant. Journal of Agricultural Technology 1 (1) : 153-162.
Three strains of the White Button Mushroom (Agaricus bisporus) were studied. First, spawn was prepared from wheat seeds covered with mycelia, and then placed within polycarbonate vials for freezing in liquid nitrogen. The effect of adding a cryoprotective solution before freezing was evaluated as a function of mycelial growth and percent viability. Two treatments were undertaken: 1) freezing with a glycerol-based cryoprotectant and 2) freezing without a cryoprotectant. Samples were maintained frozen for two weeks, after which time they were thawed and the seeds placed in Petri dishes containing a culture medium. A recovery rate of 96% was obtained for all combined samples in the experiment, whereas 95.8% of the samples frozen without a cryoprotectant were recovered. These results suggest that mycelia can survive frozen storage without cryoprotectants, provided that they are embedded within and protected by the wheat seeds in the spawn. In addition, no significant differences were observed in recovery rates and mycelial diameters between the cryoprotection and non-cryoprotection treatments when a new series of spawn was prepared from the originally-frozen mycelia.

Key words: Agaricus bisporus , edible mushroom cultivation, germplasm conservation, liquid nitrogen, white button mushroom

* Corresponding author: G. Mata;e-mail: mata@ecologia.edu.mx

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Arbuscular mycorrhizas in the Larrea divaricata scrubland of the arid “Chaco”, Central Argentina

M.A. Lugo 1, A.M. Anton 2 and M.N. Cabello 3 *

1 Área de Ecología, Facultad de Química, Bioquímica y Farmacia, Universidad Nacional de San Luis. Ejercito de Los Andes 1148, D5700HHW, San Luis, Argentina.
2 Instituto Multidisciplinario de Biología Vegetal (IMBIV – CONICET), Museo Botánico, Universidad Nacional de Córdoba, CC 491, 5000 Córdoba, Argentina.
3 Instituto Spegazzini, Facultad de Ciencias Naturales y Museo, Universidad Nacional de La Plata. Aven 53 No 477, B1900AVJ, La Plata, Argentina

Lugo , M.A., Anton, A.M. and Cabello, M.N. (2005). Arbuscular mycorrhizas in the Larrea divaricata scrubland of the arid “ Chaco ”, Central Argentina. Journal of Agricultural Technology 1 (1) : 163-178.
Arbuscular mycorrhizal root colonization and spore diversity were analyzed, in terms of seasons, dry and wet period and host species, in an arid secondary scrubland ecosystem (“Jarillal”) in Central Argentina . Larrea divaricata, dominant in this plant community, was the most colonized species followed by the grass Trichloris crinita (“herbaceous stratum fitness indicator”) whereas the lowest values werefound in Sporoboluspyramidatus and Neobouteloua lophostachya (both of them “disturbance indicators”). Root colonization was closely related to host role in the community. The AMF diversity was low because of the disturbed features of “Jarillal” and arid conditions. Spore total density, specific spore density and spore richness were more related to the seasons and water availability than to the host species.

Key words : AM colonization, AM fungal spore diversity, bushland, grassland, seasonality, soil moisture.

* Corresponding author: M.N. Cabello;e-mail: mcabello@netverk.com.armcabello@netverk.com.ar

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A simple approach to improve plant regeneration from callus culture of S orghum bicolor for crop improvement

P. Baskaran, B. Raja Rajeswari and N. Jayabalan *

Department of Plant Science, School of Life Sciences , Bharathidasan University , Tiruchirappalli -620 024, Tamil Nadu, India

Baskaran, P., Raja Rajeswari, B. and Jayabalan, N. (2005). A simple approach to improve plant regeneration from callus culture of S orghum bicolor (L.) Moench for crop improvement. Journal of Agricultural Technology 1 (1) : 179-192.
This study was conducted to establish and optimise a regeneration system for agronomically important Indian sorghum genotypes including two commercial cultivars (NSH27 and K8) of Sorghum bicolor. Callus induction and plant regeneration were achieved on transverse thin cell layers (tTCL) of hypocotyls from aseptically germinated seedlings of seven-day-old seedlings. Callus response depended on the genotype, the concentrations and composition of growth substances and number of in vitro regeneration cycles undergone by the donor plant. Murashige and Skoog (MS) medium supplemented with 4.5-18.1 µM 2, 4-Dichlorophenoxy acetic acid (2,4-D), 5.4-21.5 µM Naphthalene acetic acid (NAA), 5.7-22.8 µM Indole acetic acid (IAA) and 4.9-19.7 µM Indole butyric acid (IBA) and combined with 10% (v/v) coconut water (CW) for callus induction. The calli were cultured on MS medium supplemented with 2.2-17.8 µM 6-Benzyl aminopurine (BAP) combined with 5% (v/v) CW and addition of 2.3 m M 2, 4-Dichlorophenoxy acetic acid (2, 4-D) or 2.7 µM Naphthalene acetic acid (NAA). Highly efficient differentiations of multiple shoot buds were initiated within four weeks after inoculation. Root induction was achieved on half strength MS medium containing IAA (2.9-28.5 µM). Rooted plants were successfully acclimatized and with the survival rate reaching almost 80%. These plants grew normally without showing any morphological variation.

Key words: callus induction, coconut water, coleoptile, hardened plant, ms medium, plant growth regulators, regeneration and rooting

Abbreviations: BAP – 6-benzyl amino purine, 2, 4-D – 2, 4-Dichlorophenoxy acetic acid, CW – coconut water, IAA – indole-3-acetic acid, IBA – indole-3-butyric acid, Kin – Kinetin, NAA - a -naphthalene acetic acid.

* Corresponding author: N. Jayabalan; e-mail: jayablan54@yahoo.co.in

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