Research Journal of Biotechnology

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Role of Single Fungal Isolates and Consortia as Plant Growth Promoters under Saline Conditions

Pandya Urja and Saraf Meenu

Seven fungal isolates were screened from soil samples of different regions of Khambat and Bhavnagar. They were Aspergillus sp. (SR2, S11, N3, N4 and S17), Penicillium sp. (S12B), Fusarium sp. (S217). At 2% NaCl concentration, all fungal isolates had maximum growth after 72 h of incubation. Seven fungal isolates showed zone of phosphate solubilization on agar ranging from 15 to 30 mm within 72 h of incubation. Siderophore production was studied after 15 d of incubation on succinic acid medium. Maximum zone of siderophore production showed by isolate SR2 was 63 mm followed by isolate S17. Pot study was carried out to observe the effect of fungal inoculation on chickpea plant. Inoculation with all the fungal isolates resulted in increase in plant height, number of leaves, number of lateral roots, fresh weight, dry weight and total chlorophyll as compared to control. Penicillium sp. (S12B) and Aspergillus sp. (S11) showed increase in plant growth compared to other fungal isolates in 2% saline soils. S11 and S17 were biocompatible and showed maximum increase in all vegetative parameters compared to single culture inoculation and control suggesting that the combination could be more effective than individual strains at 2% level of salinity.

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Effect of Tillage Systems on Glomalin-Related Soil Protein in an Aquic Brown Soil

Liang W.J.

Conservation agriculture such as no tillage may increase soil carbon sequestration. The main objective of this study was to examine the profile distribution of soil organic carbon (SOC) and glomalin-related soil protein (GRSP) under no-tillage (NT), conventional tillage (CT, mouldboard ploughing) and fallow (FF, unmanaged field lasting for 6 years grown with weeds) treatments to a 1 m depth based on a 6-year tillage experiment in Northeast China. The results showed that NT enhanced SOC concentration only at the surface depths of 0–5 and 5–15 cm, but not at deeper depths. SOC concentration at the top 0–5 cm depth was significantly higher in FF than in NT treatments, but significantly lower at the 5–15 and 15–30 cm depths. Compared to CT, NT significanlly enhanced the concentrations of TG and EEG throughout the soil profile. In addition, SOC was positively correlated with TG and EEG in present study. Our study indicates that GRSP may be a useful indicator for distiguishing tillage systems on soil carbon sequestration across soil horizons.

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Impact of Cultivation Conditions on Type of Polyhydroxyalkanoate Biopolyester Production

Kanchanasuta Suwimon, Boonyawanich Sirione and Pisutpaisal Nipon

The production of polyhydroxyalkanoate biopo­l­yester from three bacterial strains, Alcaligenes euthophus, Alcaligenes faecalis and Pseudomonas oleovorans was investigated. The effect of cultivation and operational conditions; pH, temperature, nitrog­en content, aeration condition and type of co-substr­ate on the yield and type of polyester produced were examined. A. euthophus yields the maximal extent of the biopolyester under the aerobic cultivation with pH 5 and 30o C when glucose was used as a main and sole carbon source. Structural analysis using 1H NMR indicated that the accumulated biopolyester was poly (3-hydroxybutyrate-co-3-hydroxyvalerate) when pro­­pionic acid was used as co-substrate whereas poly (3-hydroxybutyrate-co-4-hydroxyutyrate) was synth­esized when acetate or butyrate were used as a co-substrate.

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Effect of C/N Ratio and Physicochemical Conditions on the production of Rhamnolipids by Pseudomonas Aeruginosa LBI

Lovaglio R. B., Costa S. G. V. A. O., Lima C.J.B., Cortezi M. and Contiero Jonas

Waste utilization for biosurfactant production is an important approach contributing to the reduction of waste treatment costs, while increasing the economic value of residues and reducing biosurfactant production costs. In an attempt to optimize rhamnolipid production by Pseudomonas aeruginosa LBI, the effects of the oil refinery waste as carbon source, C/N ratio, agitation speed, aeration rate, pH control and fed-batch culture conditions on the production of this biosurfactant were assessed in a bioreactor. Maximum production (16.9 g L-1) was attained at a C/N ratio of 23, agitation at 800 rpm and aeration at 2.0 vvm. The rhamnolipid solution (0.1%, w/v) exhibited surface and interface tensions of 25 mN/m and 0.63 mN/m respectively. The critical micelle concentration (CMC) was 73.9 mg L-1. The optimal parameters used in this study promoted the formation of a useful biosurfactant with potential for many industrial applications.

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Antibacterial Activity of Aleurites Moluccana (Euphorbiaceae) against Some Clinical Isolates

Samah Othman Abd and Razar Rasyidah Mohamad

Aleurites moluccana extracts were found to inhibit the growth of Staphylococcus aureus and Pseudomonas aeruginosa in a previous study.11 The antibacterial activities of acetonitrile, methanol and water extracts of husk, leaf and stem bark of A. moluccana were further investigated against 8 clinical isolates. The disc diffusion assays showed that crude extract of A. moluccana inhibited the growth of Staphylococcus aureus (S 277), Streptococcus pyogenes (S 526), Pseudomonas aeruginosa (P 84) and Klebsiella pneumoniae (K 6). The lowest MIC for P. aeruginosa (P 84) was exhibited by methanol extract of stem bark at concentration 98 µg/ml and S. pyogenes (S 526) at 781 µg/ml whilst both S. aureus (S 277) and K. pneumoniae (K 6) exhibited in acetonitrile of husk at 195 µg/ml and 6.25 µg/ml, respectively. The antibacterial action was bactericidal for all bacteria except for S. pyogenes (S 526) where the action was bacteriostatic. The highest polyphenols content was observed in methanol extract of stem bark. The overall results indicate that the extracts from the stem bark are potential source of phytomedicine agents for both Gram-positive and Gram-negative bacteria.

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Characterization of Commercial Microbial Products by Polymorphic DNA Markers and Enzymatic Activity Diversity: Occurrence and Potential Effects on Freshwater Mussels exposed to Municipal Effluents

Douville M., Gagné F., Zhu B., Fortier M. and Fournier M.

The increasing commercial applications of microbial products and consortia have raised concerns about their release to the environment and the potential for toxic effects on aquatic biota. This study characterized 25 bacteria, one yeast and one fungus using a combination of random amplified polymorphic DNA and 19 hydrolytic enzyme activities, with the purpose of determining their occurrence and potential effects on the immune systems of mussels exposed to a primary-treated municipal effluent for seven weeks. The amplification product (552-bp) of cpn60 gene by using universal primers was sequenced for each micro-organism and available in GenBank. The microbial community structures in the digestive gland of mussels were also characterized using 16s ribosomal gene analysis by denaturing gradient gel electrophoresis (DGGE). The results revealed that each micro-organism was easily cultivated with usual growth media and the enzyme activity profiles permitted to propose an identification method using hierarchical tree and backpropagation neural networks classifications. The bacterial community structure based on DGGE of 16S rDNA in mussels changed with the concentration of the municipal effluent. This was also revealed by a characteristic change in enzyme activity profiles with increased frequency in acid and amide phosphatase, leucine aminopeptidase and β-galactosidase in the isolated heterotrophic colonies in mussels. The analysis of heterotrophic bacteria colonies in unexposed mussels revealed that most colonies (6/10) were of unkown origin and only three bacterial colonies shared close similarities with B. subtilisis 11685, E. aerogenes, P. putida 12633. In mussels exposed to the municipal effluents, about half of the colonies were classified as unknowns while the other half was composed of bacteria showing close similarites with E. aerogenes, P. putida 12633 and 31800. These bacteria differed from those found in forming biofilms which was mainly associated to B. subtilis 11685 or P. fluorescens 13525 functional properties. The immune system of the mussels was also compromised with decreased hemocyte density, increased phagocytosis activity and cytotoxic activity. While no trend between phagocytosis activity and the enzyme activity profiles was found, the increased frequency in leucine aminopeptidase and acid phosphatase were negatively correlated with the hemocyte density and cytotoxic activity. In conclusion, a methodology to track for the occurrence, bioavailability and potential immunotoxic effects of commercial microorganisms is proposed based on a combination of genetic and phenotypic (enzyme activity) markers and immunocompetence assessment. Preliminary results indicate mussels contained only three bacteria out of 25 bacteria that bear high functional similarity and the characteristic pertubations with the enzyme activity profiles were associated with immunocompetence changes.

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Role of Chaperon Co-Expression System in the Production of Recombinant Unnatural Proteins in Escherichia Coli

Ayyadurai Niraikulam , Yoon Hyungdon and Lee Sun-Gu

Here we evaluated the chaperon co-expression system for the production of unnatural recombinant proteins through selective pressure incorporation. In this study green fluorescent protein (GFP) was utilized as model proteins and L-homopropargylglycine (Hpg) was used as a model unnatural amino acid which is a methionine surrogate and can be incorporated into the methionine residues of recombinant protein. Initial experiments showed yields of L-Hpg containing GFP proteins were almost negligible compared to those of canonical amino acid incorporated natural proteins. These results indicated that incorporation of unnatural amino acid into protein affects the normal protein folding which immediately affects the protein functionality and productivity. Co-expression of the chaperons has been well reported to enhance the protein folding and preventing the aggregation from harsh environment condition. Hence, we expected that chaperon co-expression system will facilitate the production of unnatural recombinant protein production. Coexpression of DnaK-DnaJ-GrpE chaperons systems shows that the unnatural amino acid protein production was slightly higher than the wild type. However, chaperon systems GroEL and GroES completely failed to rescue the unnatural amino containing protein so the production yield of the unnatural protein was not improved. These results demonstrated that incorporation of unnatural amino acid tremendously influences the production yield of unnatural containing proteins in Escherichia coli. As well as chaperon teams are not critically involved in protein folding of unnatural amino acid containing protein and they failed to assist stability and prevent the disaggregation of proteins.

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Growth Responses of Calotropis Procera R.Br. - A Medicinal Plant exposed to coal-smoke pollution

Kumar Sanjeev *1 and Narula Alka 2

The present work was carried out to assess the impact of coal-smoke pollution (fly ash) on growth response of Calotropis procera, a shrub of family Asclepiadaceae growing in the vicinity of coal based Badarpur Thermal Power Plant, which releases a mixture of pollutants like SO2, NO2, CO2, HF, fly ash and particulate matter. Plants growing in close vicinity with highly polluted environment showed changes in physio-chemical characteristics like incr­ease in shoot length, leaf area, biomass but consi­derable decline in chlorophyll and protein content, nitrate reductase activity, rate of photos­yn­thesis etc. Visible symptoms on the foliage were chlor­­osis and necrosis coupled with reduced growth and yield followed by premature death of plants in extreme cases.

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A Study by Absorption Spectrophotometry of the Combined Effect of Temperature and Copper Sulphate on Bilirubin Concentrations

Pillay Avin E., Salih Fadhil M. and Maleek Muthana I.

Dilute solutions of unconjugated bilirubin in aqueous NaOH were spiked with trace levels of CuSO4. The dependence of the specimen on the level of the spike and on concomitant increases in temperature was closely examined by high resolution absorption spectrophotometry. We found that the characteristic absorbance (A) of bilirubin, in the region of 453 nm dropped systematically in the presence of 1-25 mM CuSO4. This depletion was accompanied by a corresponding growth of biliverdin around 365 nm. It was discovered that the influence of temperature on the spiked solutions intensified the phenomenon. The absorbances of the species were investigated as a function of CuSO4 concentration and temperature in the range 25 – 65 oC. Appropriate statistical treatment of the data provided a means of indirectly estimating the variation in bioanalytical concentration. Consideration of ΔA vs ΔT and ΔA vs Δ [CuSO4] revealed that the thermal effect competes with the “copper sulphate effect.”

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Technology Development for Ethanol Production from the Wild Fruits of Mimusops Hexandra

Daripkar Nilambari and Jadhav B. L.*

Present investigation was carried out to develop technology for the ethanol production from the wild fruits of Mimusops hexandra using two different yeast strains. The wild fruits used in the study were collected from different parts of Maharashtra. The chemical analysis of fresh fruit showed protein, lipid, total carbohydrates and moisture content about 3.53%, 2.6%, 22% and 71.87% respectively. The fermentation of the pulp of fresh fruit was carried out by commercial MTCC – 173 strain and wild strain of S. cerevisiae. The ethanol content of the ferment was determined by Gas Chromatography. The MTCC – 173 strain produced 21% ethanol while it was 7.05% by wild strain.

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Detection of Low Molecular Weight Protein Biomarker by Serum Protein Profiling of Acute Myeloid Leukemic Patients

Sharma Chandra Kant and Sharma Vinay

Mostly cancer cells contain genetic damage for event leading to tumor genesis. The genetic damage present in a parental tumorigenic cell is maintained (i.e. not corr­ectable) such that it is a heritable trait of all cells of subsequent generations. One of the causes of gen­etic damage found in cancer cells is due to Proto-onco­ge­n­es, a protein capable of cellular transfo­rmation. The process of activation of Proto-oncogenes to oncoge­nes can include retroviral transduction or retroviral integration, point mutations, insertion mutati­ons, gene amplification, chromosomal translocation and/or pro­te­in-protein interactions. These proteins are gener­ally absent in healthy persons. These anomalous pro­te­ins may be growth factor, growth factor receptor, intra-cellular signaling molecule or enzyme of diverse signaling pathways and can be taken as biomarker for the identification of cancer in the vicinity of the begin­ing stages so that it can be treated appro­priately and cured utterly. In our study the molecular weight of the extra protein in AML patients is approximately 16.21 kDa. This abnormal serum protein can be utilized as biomarker for the early detection of AML

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Direct Shoot Organogenesis of Indian Cotton G. Arboreum CV PA 402 from Embryonic Axis Explants

Talegaokar A. P. and Dangat S.S.

Cotton is multipurpose crop serving as an engine of economic growth in both developing and developed countries across six continents. India is the only country to grow all four cultivated (both diploid as well as tetraploid) species on commercial scale. The diploid cotton species can serve as valuable gene pool for the agronomically desirable tetraploid cultivars and offer better opportunities to study gene structure and function through gene knockouts. Develo­pment of transgenic lines is one of the ways for value addition in terms of transfer of different agronomic traits as well as study of different gene functions using different tetraploid and diploid germplasm backgrounds. Regeneration protocol through somatic embryogenesis is primary requisite to exploit valuable traits available through plant transformation. In present investigation successful direct shoot organogenesis was attempted in PA 402 variety of diploid cotton using MS media supple­mented with Myo-inositol 100 mg/L Thiamine 10 mg /L Glucose 30 gm/L and agar 7 gm/L .The hormone kinetin was used in the range of 0.1 –2.5 mg/L in five different levels. Growth of shoots on MS medium supplemented with Myo-inositol 100 mg/L thiamine 10 mg /L Glucose 30 g/L and NAA 0.1 mg/L resulted in regeneration of plantlets. Among different levels of kinetin 0.1 mg/L was found the most suitable concentration for direct shoot organogenesis from embryonic Axis explants.

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Review Paper: The Utilisation of Microbial Poly-Hydroxy Alkanoates (PHA) in Food Industry

Arzu Akpinar-Bayizit, Tulay Ozcan, Oya Irmak Sahin and Lutfiye Yilmaz-Ersan

Biodegradability is defined as the capacity of to be broken down by the action of living things, such as bacteria and fungi being the main participants in the process of biodegradation in the natural world. The breakdown of materials provides them with precursors for cell components and energy for energy-requiring processes. Biodegradation is thus defined as catabolism. Some synthetic polymers, having chemical compositions resisting enzymatic attacks, can be microbially degraded with slower processes. Bioplastics is a term used for packaging materials derived from renewable resources and are considered safe to be used in food applications. In comparison to conventional plastics, the monomer composition of bio-based polymers can be differed by varying the producing strains, substrates and cosubstrates, and biopolymers such as polyhydro­xyalkanoates (PHA), polylactic acid (PLA), polysaccahrides. PHAs are carbon-energy storage material accumulated intracellularly in a variety of micro­organisms under controlled concentrations. Their biodegradability and possibility to be synthesised from renewable agricultural or even industrial wastes, make PHAs of considerable interest. Their main applications in food industry are serving as waterproof layer for the production of disposable food containers and utensils, cast films and cast sheets for thermoforming and utilisation as good odour and oxygen barriers.

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