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I take the privilege to present the hard copy compilation for the Volume 4 Issue (1) of Research & Reviews: A Journal of Biotechnology (RRJoBT). The intension of RRJoBT is to create an atmosphere that stimulates creativeness, research and growth in the area of Biotechnology. The development and growth of the mankind is the consequence of brilliant Research done by eminent Scientists and Engineers in every field. RRJoBT provides an outlet for Research findings and reviews in areas of Biotechnology found to be relevant for National and International recent developments & research initiative. The aim and scope of the Journal is to provide an academic medium and an important reference for the advancement and dissemination of Research results that support high level learning, teaching and research in the domain of Biotechnology. Finally, I express my sincere gratitude and thanks to our Editorial/ Reviewer board and Authors for their continued support and invaluable contributions and suggestions in the form of authoring writeups/ reviewing and providing constructive comments for the advancement of the journals. With regards to their due continuous support and co-operation, we have been able to publish quality Research/Reviews findings for our customers base. I hope you will enjoy reading this issue and we welcome your feedback on any aspect of the Journal.

Dr. Archana Mehrotra Director STM Journals

Research & Reviews : A Journal of Biotechnology

Contents

1. Production and Characterization of Thermostable Amylase Produced from Spore Former Thermophilic Bacilli Isolated and Screened from Teff (Eragrostis teff) Starch Rich Soil Berhanu Tsehai Andualem

2. Study of Seed Germination and Protein Content of Four Varieties of Glycine max L. â&#x20AC;&#x201C; A Miracle Crop Neha Gupta, Monika Shukla Bajpai, Pankaj K Mishra, Rita Singh Majumdar

3. Insecticidal Activity of Rotenone in Brebra (Millettia Ferruginea) Oil Refinery Residue against Different Stored Crop Insect Pests and its Prospect to Apply as Pest Control Substance Berhanu Tsehai Andualem

4. Designer Zinc Finger Nuclease: A Versatile Tool for Genome Editing in Animals Kiran Lakkireddy, Sudhakar Podha, Krishna Satya A, Kasturi K, Sambasiva Rao KRS

5. Cytotoxicity of Titanium Dioxide Nanoparticles on A Bacterial Population Isolated From Lakewater Sediments Kavyaa Venkat, Jyoti Kumari, Chandraprabha M.N, Amitava Mukherjee

6. Forward Osmosis Membrane Concentration Process of Anthocyanins from Pomegranate Fruit Chetan A. Nayak

7. Production and Characterization of Biosurfactant from Indigenous Bacterium Bacillus amyloliquefaciens USTBb: Feasibility in Microbial Enhanced Oil Recovery Radhika Chandankere, Jun Yao, Kanaji Masakorala, A.K. Jain, Ranjan Kumar

Research & Reviews: A Journal of Biotechnology ISSN: 2347-7245 Volume 4, Issue 1 www.stmjournals.com

Production and Characterization of Thermostable Amylase Produced from Spore Former Thermophilic Bacilli Isolated and Screened from Teff (Eragrostis teff) Starch Rich Soil Berhanu Tsehai Andualem* Department of Biotechnology, University of Gondar, Ethiopia

Abstract There is a need to isolate and screen amylase-producing bacilli bacteria from soil rich in content of teff starch at high temperature environment. Therefore, the aim of this study was to isolate and screen amylase-producing bacilli isolates and characterization of their optimal amylase activities. Amylase-producing bacilli were isolated and their enzymes characterized. Effect of temperature, pH, substrate and salt concentration were determined. All isolates have clear zone over 15 mm and hydrolyzed greater than 90% of starch after 60 h of fermentation. There was no significant (p ≥ 0.05) variation in enzyme productivity. Aamylases activities of G1 and G3 isolates reached their optimum at 60 °C but G2 was at 55 °C. Amylase from isolate G1 and G3 showed hydrolysis capacity of 96.84 and 97.62%, respectively, while G2 was 99.68% at 55 °C. Optimum enzyme activity of G1, G2 and G3 isolates was 100, 100 and 99.75% at pH 8, respectively. Thermal stability of all amylases was retained above 92%, while pH, G1 and G2 were above 86.37% but G2 was 82%. Thermophilic, alkalophilic and halophlic amylases have wide range of huge potntial for industrial applications.

Keywords: Alkalophile, Amylase, Insustial application, Isolate, Thermostable

Research & Reviews: A Journal of Biotechnology ISSN: 2347-7245 Volume 4, Issue 1 www.stmjournals.com

Study of Seed Germination and Protein Content of Four Varieties of Glycine max L. â&#x20AC;&#x201C; A Miracle Crop Neha Gupta, Monika Shukla Bajpai, Pankaj K Mishra, Rita Singh Majumdar* Department of Biotechnology, School of Engineering & Technology, Sharda University, Greater Noida-201301, India

Abstract Glycine max L. (Soybean) is an important source of high-quality proteins, fats, vitamins, minerals and other nutrients for both human beings and animals. The study was investigated on the seed germination, purity percentage and protein content of four varieties of Glycine max L. namely, JS335, JS9560, JS9305 and PK1042, collected from the Indian Agriculture Research Institute, Pusa, New Delhi (India). The experimental data showed that variety JS335 exhibited 85% germination, 100% purity, 0.51 mg/g protein content and 1.55 mg/g chlorophyll content. In addition, maximum purity percentage was also found in PK1042.

Keywords: Soybean, Glycine max, germination, protein content, soya oil

Research & Reviews: A Journal of Biotechnology ISSN: 2347-7245 Volume 4, Issue 1 www.stmjournals.com

Insecticidal Activity of Rotenone in Brebra (Millettia Ferruginea) Oil Refinery Residue against Different Stored Crop Insect Pests and its Prospect to Apply as Pest Control Substance Berhanu Tsehai Andualem* Department of Biotechnology, University of Gondar, Ethiopia

Abstract Use of botanical insecticides against stored crop insect pests is environmental safer and eco-friend alternative method for crop protection. The objective was to investigate insecticidal potential of oil refinery residue with rotenone against selected insect pest of stored crops. The amount of rotenone from samples was determined by HPLC. The toxicity of the rotenone from residual extract and non-defatted sample were tested against. Acanthoscelides obtectus, Sitophilus zeamais and Tribolium castaneum using standard methods. 1, 1.5 and 2 ml residue contains 1037.089, 1555.98 and 2075.57 µg rotenone, respectively. Significant differences (P ≤ 0.01) were observed among different doses of treatment for % mortality of Tribolium castanium and Sitophilus zeamais after 24 h of treatment but, there was no significant difference (P ≤ 0.01) at doses of 2 ml of residues against Tribolium castanium and 1.5 ml against Sitophilus zeamais after treatment of 24 h. Tribolium castanium seems to be more resistant than Sitophilus zeamais and Acanthoscelides obtectus for rotenone from oil refinery residue. Any way 2 ml of oil residue has potential to kill 100% of tested pests after 24 h of treatment. Therefore, the toxicity of the different extracts to tested insects was dose dependent, however, the mortality as compared to control was significant (P<0.001). So, residue from refined oil has huge potential of insecticidal activity and can serve as insect pesticide at high concentration level. It is economically important for insect crop pest control in one hand and save the money spent for inorganic insect pest chemicals and thereby reduces environmental impact of non-degradable chemicals.

Keywords: Insecticides, insect pests, Millettia ferruginea, refinery residue, rotenone, stored crop

Page 15

Research & Reviews : A Journal of Biotechnology ISSN: 2347-7245 Volume 4, Issue 1 www.stmjournals.com

Designer Zinc Finger Nuclease: A Versatile Tool for Genome Editing in Animals Kiran Lakkireddy*, Sudhakar Podha, Krishna Satya A, Kasturi K, Sambasiva Rao KRS Department of Biotechnology, Acharya Nagarjuna University, Nagarjuna Nagar-522510, Guntur, Andhra Pradesh, India

Abstract In this review, is provided an update on the most popular approaches, including zinc finger nuclease (ZFN)-based gene editing by homologous recombination (HR), gene knockout, non-homologous end joining (NHEJ), and gene therapy in mouse. Zinc finger nucleases (ZFNs) are synthetic restriction enzymes evolved by fusing zinc finger that facilitate genome editing by creating a double-stranded break in DNA at a user-specified location. ZFNs are new tools that promise to radically simplify gene knockout, targeted gene deletions and replacements, because of their high specificities. Most recent strategies for developing ZFNs with customized sequence specificities require the construction of numerous tandem arrays of zinc finger proteins (ZFPs) to target genes specifically, also added more fingers (up to six per ZFN) to specify longer and rare cleavage site. The classical synthetic Cys2His2 zinc finger domains (C2H2 ZFs) with engineered DNA-invoking specificities have shown promise for application in biological research. Current methods of an engineered ZFN design, the important use in modification of the mouse genome, and innovative applications of this technology in mouse will be discussed. Targeted genome inactivation of endogenous immunoglobulin genes in combination with addition of human immunoglobulin genes has been achieved in mice. A light-activated ZFN will competence the conditional generation of gene knockins and knock-outs with unique accuracy. ZFNs are useful to generate precisely targeted genome edits resulting in cell lines including somatic cell lines with targeted gene deletions, integrations or modifications. ZFN-mediated site-specific mutagenesis have been proven to be effective in many model organisms â&#x20AC;&#x201C; arabidopsis, drosophila, zebrafish, caenorhabditis elegans, rabbit and mouse.

Keywords: Zinc finger nuclease, double-strand base pairs, gene targeting, homologous recombination, non-homologous end joining, gene therapy, animal models

Research & Reviews: A Journal of Biotechnology ISSN: 2347-7245 Volume 4, Issue 1 www.stmjournals.com

Cytotoxicity of Titanium Dioxide Nanoparticles on A Bacterial Population Isolated From Lakewater Sediments Kavyaa Venkat1, Jyoti Kumari2, Chandraprabha M.N.1*, Amitava Mukherjee2 1

Department of Biotechnology, M.S. Ramaiah Institute of Technology, Bangalore, India 2 Centre for Nanobiotechnology, VIT University, Vellore, India

Abstract Studies suggest that approximately 4.6 million ton of pigmentary titanium dioxide (TiO2) is consumed annually worldwide. Owing to the increased industrial and consumer applications, there is growing concern regarding the possible risk associated with their environmental exposure. TiO2 dust, when inhaled, has been classified by the International Agency for Research on Cancer (IARC) as an IARC Group 2B carcinogen, meaning it is possibly carcinogenic to humans. TiO2 nanoparticles (NPs) can enter the aquatic environment either through direct or indirect release as effluents from nano-paints, sunscreen lotions, food additives, medical use, dismantling of batteries, etc. Therefore, the aim of the present study was to explore the difference in toxicity mechanism of TiO2 NPs at low concentrations (1 Îźg/ml), in a bacterial consortium consisting of Pseudomonas aeruginosa, Bacillus altitudinis and Bacillus tequilensis, isolated from lakewater sediments, under light and dark conditions. The cytotoxicity of the NPs was studied by performing several biochemical assays for the assessment of membrane permeability, cell viability, oxidative stress and secretion of exopolysaccharides. It was found that there was generation of ROS, which may have resulted in the decrease in cell viability. Also, the cytotoxic effects were found to be more pronounced under dark than under light conditions.

Keywords: titanium dioxide nanoparticles, cytotoxicity, ROS

Research & Reviews: A Journal of Biotechnology ISSN: 2347-7245 Volume 4, Issue 1 www.stmjournals.com

Forward Osmosis Membrane Concentration Process of Anthocyanins from Pomegranate Fruit Chetan A. Nayak* Department of Chemical Engineering, BMS College of Engineering, Bangalore, India Abstract The production of concentrated pomegranate juice was investigated by using forward osmosis membrane concentration process. Suspended solids were completely removed in the clarification step, while soluble solids and organic acids were concentrated using forward osmosis membrane process. The processes was performed at ambient temperature (25 ± 2 °C) and non pressure driven process to produce a concentrated extract to a total soluble solids (TSS) content of 60 Brix from an initial concentration of 4 Brix (15 times increase). The antioxidant activity of pomegranate aril juice, attributed to a great extent to total phenols and anthocyanins content, was efficiently preserved during the concentration step independently on the achieved level of total soluble solids. Non-enzymatic browning index and degradation constant for thermally concentrated sample were found to be approximately two (0.78–0.35) and eight (63.0×10−3 to 8.0×10−3 day−1) times, respectively, as compared to the concentrate produced by forward osmosis. An integrated membrane process scheme for the production of concentrated pomegranate juice with potential applications for food, pharmaceutical and cosmetic sectors was proposed.

Keywords: forward osmosis, pomegranate, membrane process, anthocyanins pigment

Page 36

Research & Reviews: A Journal of Biotechnology ISSN: 2347-7245 Volume 4, Issue 1 www.stmjournals.com

Production and Characterization of Biosurfactant from Indigenous Bacterium Bacillus amyloliquefaciens USTBb: Feasibility in Microbial Enhanced Oil Recovery Radhika Chandankere1, Jun Yao1,2*, Kanaji Masakorala1,3, A.K. Jain4, Ranjan Kumar5 School of Civil & Environmental Engineering and National “International Cooperation Based on Environment and Energy”, and Key Laboratory of “Metal and Mine Efficiently Exploiting and Safety” Ministry of Education, University of Science and Technology Beijing, 100083 Beijing, PR China 2 Key Laboratory of Biogeology and Environmental Geology of Chinese Ministry of Education, and Sino-Hungarian Joint Laboratory of Environmental Science and Health, University of Geoscience, Wuhan 430074, PR China 3 Department of Botany, Faculty of Science, University of Ruhuna, Matara, Sri Lanka 4 Domain Expert, Petroleum Microbiology and Unconventional Oil/Gas, Oil and Natural Gas Corporation Ltd. , Dehradun, India 5 Department of Civil Engineering, Indian Institute of Technology Roorkee, Uttarakhand, India 1

Abstract In the present study, the production and characterization of biosurfactant by an indigenous Bacillus methylotrophicus USTBb (GenBank accession no. KF496216) was investigated. This bacterial strain was isolated from crude oil contaminated aqueous medium was identified to be a persuasive producer of biosurfactant using crude oil as the sole carbon and energy source. Growth parameters were optimized, with the aim of increasing biosurfactant productivity. Maximum biosurfactant production (5.5 g L-1) by strain USTBb was reached after 12 d of incubation time and was found to be growthassociated. The strain was able to reduce the surface tension of its cell free culture supernatant containing biosurfactant to 28 mN m-1. Biosurfactant exhibited excellent emulsification activity against crude oil when compared with various tested hydrocarbon and vegetable oil and it possessed a low critical micelle concentration (CMC) value of approximately 35 mg L-1, indicating high surface activity. USTBb biosurfactant showed stability under extreme environmental conditions such as high temperature, pH and salinity. Elemental compositions and thermal analysis of biosurfactant was determined by X-ray photoelectron spectroscopy (XPS) and thermal gravimetric (TG) respectively. Furthermore, biosurfactant was found to be an antimicrobial agent against gram positive and Gram negative bacterial strains. Studies on the crude oil recovery using biosurfactant produced by Bacillus sp. USTBb suggested its prospective relevance in microbial enhanced oil recovery (MEOR) and bioremediation.

Keywords: biosurfactant; surface tension; crude oil; microbial enhanced oil recovery; stability; characterization