Scientific Program

Conference Series Ltd invites all the participants across the globe to attend 13th Biotechnology Congress San Francisco, USA.

Day 3 :

  • Poster Presentation
Location: Sausalito
Biography:

Dr. Aka-Kacar has received her MSc. (1994) on plant tissue culture and PhD. (2001) degree on molecular characterization at Horticulture Department of Çukurova University in Adana,Turkey. During her PhD period she studied in Michigan State University, Plant and Soil Science Department, USA (TUBITAK-NATO Fellowship), East Malling Research Institute, UK, Bologna University, Horticulture Department Italy, as a visiting scientist. She spent a postdoctoral period at University of California Riverside, USA on genetic mapping in Citrus (2004-2005). She received Assoc.Prof title in 2006 and she has a full Prof position since 2013 at Cukurova University, Turkey.

Abstract:

Pomegranate (Punica granatum L.) is one of the oldest known edible fruit tree species, originating in Central Asia, but with a wide geographical global distribution. Besides using pomegranate as raw fruit, it has been used as herbal remedy. In consumption of pomegranate soft-hard seededness is very important. Soft seededness arises in a reduction of lignin. Lignin topo chemistry has ultra-structural aspects and lignification results from the enzyme mediated polymerization. Also lignin has three different monomers (coniferyl alcohol, sinapyl alcohol, p-coumaryl alcohol) are synthesized in the cytoplasm. Aim of the present study is to determine initiation time of lignification after pollination and genes involved in lignification mechanism in soft and hard seeded pomegranates. Known as a hard-seeded Hicaznar and soft-seeded 33N26 varieties were used as plant materials. Fruits from the two defined varieties were taken at intervals after pollination and fertilization at different sizes. Seed samples were used for transcriptome sequencing. Primary sequencing were produced by Illumina HiSeqTM 2000, called as raw reads, was subjected to quality control (QC). After QC, raw reads were filtered into clean reads aligned to the reference sequences. De novo analysis was performed to detect genes expressed in seeds of pomegranate varieties. We performed downstream analysis including gene expression, deep analysis based on gene expression, deep analysis based on DEGs, including Gene Ontology (GO) enrichment analysis. This dataset provides valuable information regarding pomegranate transcriptome changes for mechanism of soft-hard seeded pomegranate and may help guide future identification and functional analysis of genes that are important for lignification.

Biography:

Wook Jin Kim is currently a Senior Research Scientist at the Korea Institute of Oriental Medicine (KIOM). He works a development of DNA marker for discriminating betweenauthentic medicinal plant species and adulterants since 2012.

Abstract:

GennusPaeonia is an important medicinal plant in Asian traditional medicines. Among Paeonia species, P. lactiflora, P. japonica, P. veitchii, and P. suffruiticosa are pharmaceutically defined in different ways in the national pharmacopieias in Korea, Japan, and China.The roots of three Paeonia species, P. lactiflora, P. japonica andP. veitchii, commonly has been used as Paeoniae Radix, and the root bark of P. suffruticosahas been used as MoutanRadicis Cortex in Korean Traditional Herbal Medicine. However, only the roots of P. lactifolora and root bark of P. suffruticosa is pharmaceutically described as Paeoniae Radix and MoutanRadicis Cortex, respectively, in the pharmacopoeia of China and Japan.Since the morphological similarities of root and aerial part of these species, the identification of accurate species is very difficult. In addition, these herbal medicines bistributed as dried root slices or processed medicinal ingredients in the herbal market. Therefore, it is important to authenticate the different species used in these herbal medicines. So, we analyzed DNA barcode seqeunce of rDNA-ITS regionusing 17 samples of four Paeoniaspecies, and thenobtained species-specific marker nucleotides that can be used as genetic markers to identify these four plants at the species levels.Based on rDNA-ITS sequences, peptide nucleic acids (PNA) analysis which of probe-based fluorescence melting curve analysis was carried out to develop a powerful technique for detecting species-specific point mutations, namely marker nucleotide or single nucleotide polymorphism, capable to discriminate the fourPaeonia species without sequence analysis. Also, this method can provide rapid and efficient authentication of four Paeoniaspecies. Therefore, PNA analysis of four herbaciousPaeonia species will help to accurately authenticate each species and standardize the origin and quality of Paeoniae Radix and MoutanRadicis Cortex.

Biography:

Birhanu KINFU studied his BSc at the University of Gondar in Applied Biology and his MSc at Addis Ababa University in Biotechnology. He completed  both degrees with great distinction. He has worked as university lecturer for 2 years. Winning the prestigious DAAD research grant award  (German academic exchange service) under its ‘young academics and scientists’ program,  he is currently doing his PhD at the University of Hamburg, Germany, Microbiology and Biotechnology department. His main work focuses on biocatalytic phosphorylation of selected metabolites,  metagenomics and in vitro protein expression systems with manuscripts on pipeline.

Abstract:

D-Glycerate-2-phosphate is an important substrate and crucial metabolite of central carbon metabolism, glycolysis/ gluconeogenesis and pentose phosphate pathway, glycine, serine and threonine metabolism, methane metabolism, biosynthesis of plant secondary metabolites, phenylpropanoids, terpenoids and steroids, alkaloids derived from shikimate pathway, antibiotics, amino acids. Therefore an efficient, robust and scalable route for the preparation of enantio pure D-glycerate-2-phosphate is needed. A straightforward one-step biocatalytic phosphorylation of glyceric acid catalyzed by a recombinant glycerate 2-kinase heterologously expressed as maltose binding protein fusion has been investigated using racemic and the enantiopure D- and L-glycerate as substrate. The reaction was coupled with the phosphoenolpyruvate/ pyruvate-kinase-system for ATP-regeneration and monitored by 31P-NMR spectroscopy. This phosphorylation reaction using recombinant glycerate 2-kinase is highly enantio-selective and sustainable, as it yields enantiomerically pure D-Glycerate-2-phosphate in less reaction steps and with higher purity than chemical routes.

Wojciech Hahnel

Hochschule Kaiserslautern University of Applied Sciences, Germany

Title: Development of an automated portable system for extraction of DNA from difficult samples
Biography:

Wojciech Hahnel, 28 years old, has completed his B.Sc. in Biotechnology in 2011 at Mannheim University of Applied Sciences, Germany, followed by a M.Sc. in Biotechnology with focus on “Bioprocess Development” in 2013 at the same university. Since 2014, Wojciech Hahnel is doing his PhD at University of Applied Sciences Kaiserslautern, Germany, which he is planning to finish in 2017. During his studies he has been working at the Dublin City University (DCU) in Dublin, Ireland, and at the University Hospital in Bochum, Germany.

Abstract:

DNA extraction from difficult samples (e.g. stool, soil) is problematic, as there are inhibitors that influence further processing of DNA and compounds which are mutagenic and destructive against DNA. Especially fecal DNA has diagnostic relevance because it contains DNA from various sources, like blood or intestinal mucosa. Analysis of fecal DNA might allow conclusions on the presence of (intestinal) diseases like tumors and inflammations at early stages in a quick and non-invasive manner.

Kits for DNA extraction from difficult samples are commercially available, but these are neither automated nor quick nor easy to use. Thus, an easy automated portable system for extraction of DNA from difficult samples would be beneficial. It could be used on-site, e.g. bedside in hospitals.

After testing different strategies and kits, the geneMAG-RNA/DNA kit from chemicell using magnetic beads was found to be suitable. Since this kit has been developed for other sample types, it was adapted and scaled down to a portable microchip system. For further evaluations soil was used because of easier availability and handling. Centrifugation was replaced by sedimentation. As a result complete DNA extraction could be performed inside the chip. As pumping is performed by air pressure, most of the tubing is not in contact with liquids and can be reused. The chip itself can be reused as well.

Future plans include further automation of the system, testing of different sample types and evaluation of the sedimentation step. Secondly, we test pre-filled chips in order to provide a ready-to-use system.

Biography:

Araceli Olivares is a Biochemical Engineer at the Pontifical Catholic University of Valparaiso (PUCV), Dr. in Engeniering  Science m/Biochemical Engineering in PUCV. Since 20011 is a researcher of the Center for Studies on Healthy Food (CREAS). She has 3 published patents (one in Chilean and two in Unites State Patent Ofice) and 5 application patents in USPTO. She has 8 scientific journals.

Abstract:

Probiotics must be delivered alive to exert a positive health effects in site of action (the intestine). Once reach the intestine, they should establish themselves and exert a positive health effects. They must be survive the stomach acidic, bile acid and others degradative enzime througth intestinal tract.

The aim of current study was to evaluate the performace of microencapsulated Lactobacillus casei using alginate gel matrix to facilitate a suitable carrier system to enhance this objetive and confer protection from the acidity.

A solution with lyophilized L. casei at 5g/L (108 CFU/mL) was prepared and mixed (1:1) with sodium alginate solution at 2%. As a hardening solution, calcium chloride at 0.1M with gently stirring was used. Microencapsulator provide by BÜCHI (Encapsulated B-390) was used.

Simulated gastric juice (SGJ) with 9g/L of sodium chloride and 3g/L of pepsin was prepared. Different pH was adjusted with 1M HCl. 400mg of microspheres containing L. casei were mixed in 20mL of SGJ with pH adjusted at 2.0; 2.5; 3.0; 3.5 and 4.0 and incubated until 120 min a 37oC and 50rpm. Microencapsulated Lactobacillus were filtered and dissolved in sodium citrate to released and count CFU in supernatant.

The best results were obtained at pH 4.0 (over 70% of survival at 90min). At pH 3.5 the survival was over 20% at 60 min, however at pH less than 3.0 there was a rapid loss of viability and the survival was null at 90 min. Then, a microencapsulated is an available technique to maintain viability of probiotics to protect the passage through intestinal tract.

Acknowledgment: Project Fondecyt No 11130460

Biography:

Jee-Yeon Ryu is a PhD candidate from Ajou University College of Pharmacy. She is the director of Tae-Jong Yoon. She is studying synthesis of nanomateials aimed at biological applications..

Eun Yi Cho has completed her PhD from Chonnam National University, South Korea in 2011  where she majored biochemistry and then she is post-doctoral researcher at the  Ajou University College of Pharmacy through the korea research institute of chemical technolgy (KRICT). She is currently the study of gene edting using nanotechnology.

Abstract:

Gene editing (CRISPR/Cas9) technology has been spotlighted as a superior therapeutic tool that is capable of treating the fundamental causes of disease induced by genetic abnormalities, which have to date been considered as incurable diseases. Thus many researchers have focusing establishment of gene editing moiety delivering, because efficient and safe delivery in the body remains one of the major challenges of biomedical and nano-pharmaceutical research. A plasmid based CRISPR/Cas9 system has shown several critical limitations such as off-targeting, integration of DNA segment and toxicity of transfection agent. To overcome these problem, protein based CRISPR/Cas9 system was recently co-opted to the therapeutic or gene editing method. However, the protein system has still remained the stability problems, especially in in vivo system as like degradation by enzymatic reaction or low efficiency.

Herein, we elucidate novel method for the delivery system of protein based CRISPR/Cas9 with high efficient and bio-compatibility. The CRISPR/Cas9 complex was successfully encapsulated into the nanometer sized liposome (nanoliposome), which was composited with bioapplicable phospholipid chemicals through metal coordiation reaction. After chemical surface modifcation, the nanoliposome with gene editing materials was shown long term solution stability without agglomeration and penetrated well into the cell cytosol. We suggest optimization of preparation for the nanoliposomal protein based CRISPR/Cas9 system as a platform particle to the therapeutic application. As a proof of concept, the nanoliposome with CRISPR/Cas9 system was exploited to apply for the type 2 diabetes therapy and exhibited effectiveness of glucose control without off-targeting, acute toxicity and degradation. More importantly, the our particle platform system was shown higher regulation effect than a clinical used chemical drug without various adverse reactions effect including renal disorder or allergic reaction.

Biography:

The presenting author, Gourab Ghosh is currently pursuing his Ph.D. in transgenic crop science from St. Xavier’s College, Kolkata affiliated under University of Calcutta, India. He is 27 years of age. He has two publications in reputed, peer-review journals to his name.

Abstract:

Pigeonpea is one of the major grain legumes of tropics and subtropics, covering vast regions of developing countries from Africa, Asia to Latin America. It ranks fifth in area among pulses after soybean, common bean, peanut, and chickpea.Globally, pigeonpea is cultivated on 4.92 million hectares with an annual production of 3.65 metric tons and productivity of 898 kg/ha2.As they are grown in harsh environments and exposed to a variety of biotic and abiotic stresses, their pro­ductivity has not increased conspicuously for the last 50 years.Among many insect pests, the pod borer Helicoverpa armigera causes significant damage to this crop. It is the most devastating Lepidopteran pest and causes extensive economic losses to the tune of US$ 300 million annually. The present study seeks to protect pigeonpea plants from H. armigera infestation by incorporating cry1Ac and cry2Aa genes, through a unique and efficient gene transfer method. An Agrobacterium tumefaciens-mediated transformation strategy was implemented using in vitro transgenic shoot-grafting technique. A. tumefaciens harbouring different binary vectors containing cry1Ac and cry2Aa geneswere used for transgenic pigeonpea development. An overall 7-9% of transformation frequency was recorded. After monitoring transgene integration by Southern hybridization, transgenic T1 and T2 lines were further analyzed by western blot, ELISA and insect bioassay. Transgenic lines obtained,exhibited optimum expression of Cry1Ac and Cry2Aa proteins. This study was further extended to the development of selectable marker (nptII) free cry1Ac expressing transgenic lines using cre-lox mediated marker elimination system

Miyu Nishiguchi

Nara Institute of Science and Technology, Japan

Title: Ergothioneine fermentative production in Escherichia coli
Biography:

Miyu Nishiguchi is second year master’s student in NARA INSTITUTE of SCIENCE and TECHNOLOGY. She study applied microbiology.  

Abstract:

Ergothioneine (ERG) is sulfur-containing amino acid synthesized by certain bacteria and fungi. Recently, findings point to critical functions in human physiology. Human takes ERG from food and concentrates it in specific tissues or cells such as liver, kidney, central nervous system, and red blood cells. ERG is marketed as dietary supplement or nutraceutical so that acts as anti-oxidant. It has been recognized that filamentous fungi or Actinomycetes produce ERG. However, in 2010, the ERG biosynthetic gene was identified for the first time. Here, we challenged to produce ERG from glucose with our constructed cysteine producer. E. coli has a regulation system that synthesized cysteine from energetically-favored thiosulfate, as the assimilation of sulfate spends 2 ATP and 4 NADPH. This cysteine producer produces 16 g/L of cysteine from thiosulfate. Therefore, we established world-first ERG fermentation and challenged production of more cheaper ERG.

  We cloned ERG biosynthetic genes from Micobacteria smegmatis, and performed heterologous expression of cloning ERG genes in E. coli. The analysis of the culture medium by LC-MS/MS detected ERG peak. When a plasmid carrying these ERG biosynthetic genes was introduced into cysteine producer with enhanced biosynthesis, weakened degradation, and improved export of L-cysteine, the transformants slightly produced ERG into medium from thiosulfate (30mg/L of ERG). Interestingly, this transformants produced 200 mg/L of ERG from sulfate. We propose that the spend of NADPH is important for production of ERG.

(This work was supported by the Programme for Promotion of Basic and Applied Researches for Innovations in Bio-oriented Industry to I.O.)

Biography:

Rayssa Almeida Garcia is a PhD student in the Molecular Biology Program of the Federal University of Brasilia – Brazil,  at the age of 23. She does her research work at in the Plant-Pest Interaction laboratory at Embrapa Genetic Resources and Biotechnology- Brasilia/ Brazil, under supervising of Dr Maria Fatima Grossi-de-Sa, have published  a large number of papers in reputed journals and is a fellow member of the World Academy of Science.

Abstract:

Gene silencing through RNA interference as a biotechnological approach for the control of crop insect-pests have been intensively applied in the last few years. dsRNA microinjection and in vitro feeding are the most wildly used approaches for administering RNAi in insects. However, RNAi efficiency appears to be variable among different insect groups, especially when applied by feeding - for some insect groups the oral delivery of the dsRNAs has been reported highly ineffective. In initial studies, our gene silencing data for cotton boll weevil (Anthonomus grandis)  were unclear when dsRNA administration was done by feeding. The purpose of this work was to assess the possibilities of RNAi as a tool for the control of this insect pest using oral delivery of dsRNAs and to investigate the reason for the low efficiency in gene silencing,  aiming to develop a strategy to deal with the efficiency and usage of dsRNA by feeding. Data showed an optimal nucleasic activity of the A. grandis gut nucleases at acid pH, ranging from 5.5 to 6.5, and the A. grandis' gut homogenate significative degraded both dsRNA and dsDNA. Three nuclease sequences were found in A. grandis transcriptome, named AgNuc1, AgNuc2, and AgNuc3; in which AgNuc2 and AgNuc3  showed to be highly expressed in the insect gut. The silencing of the three nuclease genes strongly diminished dsRNA degradation when dsRNA were incubated with homogenate from silenced insects. On the other hand, when dsRNAs were protected with a Cell Penetrating Peptide (CPP) fused with a dsRNA Binding Domain (DRBD), no dsRNA degradation was found. Furthermore, dsRNAs complexed with CPP-DRBD were found to enter into A. grandis gut cells. The dsRNA complexed administered in the diet caused a greater gene silencing, compared to naked dsRNA. All data point out to the relevance for overcoming the gut nucleases with/or in parallel with the RNAi applications for the control of crop insect-pests.

Biography:

Eman Ramadan finished Bachelor of Medicine, Bachelor of Surgery (M.B., B.C) Ain shams university faculty of Medicine in 1997. Then finished M.Sc. Degree in pediatrics from Ain shams university faculty of Medicine 2002. In 2011, received M.Sc. in Biotechnology from American university in Cairo. Currently PhD Candidate: American university in Cairo Egypt, expected graduation fall 2016.

Abstract:

Red SeaKebrit Brine (1490m)possess unique environmental conditions, characterized by high salinity 4 M, temperature 23.4°C, elevated concentration of heavy metals, no oxygen and high hydrostatic pressure. In order to highlight the structural-functional relationship of enzymes adaptation to such extreme environmental conditions, DNA isolated from the microbial community of Kebrit Brine is subjected to 454-pyrosequencing and a metagenomic dataset is established; and looked for enzymes involved in mercury detoxifications. An operon containing the genes essential for mercury detoxification was identified in our 454-pyrosequencing metagenomic dataset.  A total of 28merAOrthologs were identified in Kebrit Brine metagenomics library, choosing two merA genes: one representing the consensus sequence (K35-NH) and the other (K09-H) has amino acid substitutions replacing non-polar with acidic amino acids. Kinetic parameters were measured at the NaClconcentration that gave maximum activity for the respective enzyme. K09-Hmaximal  activation  is  observed  at 2Molar NaCLand  retains 65% of its activity at 3 Molar. K35-NH showed maximum activity at 0 Molar NaCl which is equvlent to 52% activity of K09-H at 4 M. Both and K35-NH retained 90%of their activity after 10 minutes incubation at 65˚C.Comparing the kinetics of both enzyme suggested, that the amino acids difference between the two orthologs are acquired evolutional structural adaptation  to confer site-specific level of halophilicity to survive in such extreme environment as KebritDeep.Thishalophilic feature if used properly is a potential for many industrial and bioremediation applications in detoxification of mercury.

Biography:

Shota Kajiwara was recieved the Master of Engineering degrees in Osaka Prefecture University, Osaka, Japan, in 2016. He is now a doctoral course student of Osaka Prefecture University. His research interests includes the stabilization of enzyme in the presence of organic solvents.

Abstract:

Lipases are one of the most important enzymes as industrial catalysts. When lipases are used in the presence of organic solvents, the reaction rate is remarkably increased by solubilizing the poorly soluble substrates. Furthermore, lipases can catalyze ester synthetic reactions and transesterification reactions in the presence of organic solvents. However, enzymes including lipases are generally inactivated in the presence of organic solvents by direct contact with molecules of organic solvents. Some coordinated water molecules exist around sugars such as sucrose. Thus when lipase was complexed with sucrose and used in the presence of organic solvents, coordinated water molecules around the lipase-sucrose complex might reduce direct contact between lipase and organic solvent molecules. In this study, the transesterification activity of the lipase-sucrose complex in the presence of organic solvents was examined. The lipase-sucrose complex was prepared by freeze-dry of a lipase solution containing 1% (w/v) of sucrose (mass ratio of lipase:sucrose = 1:130). The lipase-sucrose complex showed the higher transesterification activity than freeze-dried lipase without sucrose in the presence of n-hexane and n-hexane containing 50% (v/v) of n-octane, 1-propanol, 1-pentanol, or 1-octanol. After incubation in 100% of n-hexane, n-octane, or 1-propanol at 30˚C for 24 hours, the residual transesterification activity of lipase-sucrose complex was higher than that of the freeze-dried lipase without sucrose. Furthermore, after incubation in 100% of 1-pentanol or 1-octanol, the transesterification activity of lipase-sucrose complex was also increased. In conclusion, the transesterification activity and organic solvent-stability of the lipase were successfully improved by complexation with sucrose.

Biography:

MohamadMaged has graduated from the Faculty of pharmacy, Cairo University in 2007. He has completed his MSc.in Biotechnology at the American university in Cairo (AUC) in 2011 and expected to finish his PhD from the AUC in December 2016. He is also a nutritionist and a personal trainer.

Abstract:

The lower convective layer (LCL) of the Atlantis II brine pool of the red sea is a unique environment characterized by high salinity of 4.4 Molar, temperature of 68οC and very high concentrations of heavy metals. Mercuric reductase enzymes functional in such extreme conditions could be used inthe environmental detoxification of mercurial poisoning. This would be of use in the Egyptian Gold mines, where mercuryused in the extraction process imposes a serious ecological hazard.We constructed an environmental Atlantis II mercuric reductase library, where we have identified two classes of mercuric reductases. One is the halophilic/thermostablemerA. It is designated ATII-LCL-H. It retains 50% of its activity after 10 minutes incubation at 75˚C. The other is a non-halophilic /thermostablemerAdesignated as ATII-LCL-NH. Itretains 61% of its activity at 65˚C.The ATII-LCL-HmerAhas two characteristic signature boxes and a short motif composed of 4 Aspartic acids (4D414-417). In order to understand how two enzymes from the same environment have evolved to withstand heat, we mutated the isoform ATII-LCL-NH.Substitution of 2D at positions 415/416 enhanced the thermal stability while other mutants did the opposite effect. The 2D mutant retains 88.6% of its activity at 65˚C. Three-dimensional structure prediction revealed newly formed salt-bridges and H-bonds in the 2D mutant as compared to the parent molecule.

Biography:

Abril Gomez-Guzman is currently a graduate student in the University of Guadalajara Jalisco, Mexico. She is doing her Ph.D studies in the program of Science of Biotechnological Processes. She has completed her master studies from the Autonomous University of Baja California at the age of 25 year and her bachelor degree studies from the Technological Institute of Tepic, in Biochemical Engineering. She was a professor of the Autonomous University of Guadalajara in the period of 2011-2013. She has published one paper in reputed journals and  she is working in another ones.

Abstract:

Microalgae and some bacteria offer a promising technology for the removal of nutrients such as Nitrogen and Phosphorus in municipal wastewater. In this investigation the microalgae Chlorella vulgaris, the bacteria Pseudomonas putida, Bacillus cereus and the consortium microalga/bacteria of these microorganism were used as model in the removal of nutrients using a model wastewater. The results of removal of nutrientes with Chlorella vulgaris was 24%, 80.6% and 4.30% for NO3-N, NH3-N and PO4-P, respectivley. For Bacillus cereus the removal of nutrients was 8.4%, 28.8% and 3.8% of NO3-N, NH3-N and PO4-P. With Pseudomonas putida was 5%, 41.8% and 4.3% of NO3-N, NH3-N and PO4-P (in 168 hours respectively). The consortium of Chlorella vulgaris/Bacillus cereus/Pseudomonas putida were able to remove the 29.4%, 4.2% and 0% of NO3-N, NH3-N and PO4-P in 240 hours. The highest biomass production was found with Bacillus cereus 450 mg/L followed by Pseudomonas putida 444 mg/L, the consortium 205 mg/L and Chlorella vulgaris 88.9 mg/L. This study showed that Chlorella vulgaris consume first NH3-N as a source of Nitrogen and when it is exhausted then consume NO3-N, it happen when both compounds are available in the medium. In the removal of phosphate (PO4-P) the microorganisms Chlorella vulgaris and Bacillus cereus were able to remove a higher amount because they can assimilate this nutrient even in stress conditions (like changes in pH) such as happening in the municipal wastewater.