National Institute of Genetic Engineering and Biotechnology of Iran
Iranian Journal of Biotechnology
1728-3043
2322-2921
17
3
2019
09
01
Identified Hybrid tRNA Structure Genes in Archaeal Genome
1
8
EN
Uttam
Roy Mandal
Department of Mathematics, Raidighi College, Raidighi, W.B., India
urmandal@gmail.com
Shib
Sankar Das
Department of Mathematics, Uluberia College, Uluberia, Howrah, W.B, India
ssdas80@gmail.com
Brajadulal
Chattopadhyay
Department of Physics, Jadavpur University, Kolkata, W.B, India
bdc@phys.jdvu.ac.in
Satyabrata
Sahoo
Department of Physics, Dhruba Chand Halder College, Dakshin Barasat, W.B., India
dr_s_sahoo@yahoo.com
10.29252/ijb.2254
<strong>Background:</strong> In Archaea, previous studies have revealed the presence of multiple intron-containing tRNAs and split tRNAs. The full unexpurgated analysis of archaeal tRNA genes remains a challenging task in the field of bioinformatics, because of the presence of various types of hidden tRNA genes in archaea. Here, we suggested a computational method that searched for widely separated genes encoding tRNA halves to generate suppressive variants of missing tRNAs.<br /> <strong>Objectives: </strong>The exploration of tRNA genes from a genome with varying hypotheses, among all three domain of life (eukaryotes, bacteria and archaea), has been rapidly identified in different ways in the field of bioinformatics. Like eukaryotic tRNA genes, it has been established that two separated regions of the coding sequence of a tRNA gene are essential and sufficient for promotion of transcription. Our objective is to find out the two essential regions in the genome sequence which comprises two halves of the hidden tRNAs.<br /> <strong>Material and Methods:</strong> Considering the existence of split tRNA genes widely separated throughout the genome, we developed our tRNA search algorithm to predict such separated tRNA genes by searching both a conserved terminal 5'- and 3'-motif of tRNA in agreement with the split hypothesis on the basis of cloverleaf prediction and precise insilico determination of bulge-helix-bulge secondary structure at the splice sites.<br /> <strong>Results:</strong> By a comprehensive search for all kinds of missing tRNA genes, we have constructed hybrid tRNA genes containing one essential region from tDNA (XYZ) and the other from tDNA (ABC), both from same species in the archaea. We have also found, this type of hybrid tRNA genes are identified in the different species of the archaea (XYZ: ASN, ARG and MET; ABC: ASP,SER, ARG and PRO).These hybrid split tRNA share a common structural motif called bulge-helix-bulge (BHB) a more relaxed bulge-helix loop (BHL), at the leader exon boundary and suggested to be evolutionary interrelated.<br /> <strong>Conclusions:</strong> Analysis of the complete genome sequences of <em>Metallosphaera sedula</em> DSM 5348, Desulfurococcus kamchatkensis 1221n and Ignicoccus hospitalis KIN4/I in archaea by our algorithm revealed that a number of hybrid tRNAs are constructed from different tDNAs . Asymmetric combination of 5’ and 3’ tRNA halves may have generated the diversity of tRNA molecules. Our study of hybrid tRNA genes will provide a new molecular basis for upcoming tRNA studies.
Helix (Snails),Open Reading Frames,RNA, Transfer
https://www.ijbiotech.com/article_85191.html
https://www.ijbiotech.com/article_85191_2ccf57acbb2d42dcc114bf12dcadd316.pdf
National Institute of Genetic Engineering and Biotechnology of Iran
Iranian Journal of Biotechnology
1728-3043
2322-2921
17
3
2019
09
01
RNA-Seq Bayesian Network Exploration of Immune System in Bovine
9
17
EN
Elham
Behdani
Department of Animal Science, Faculty of Animal and Food Science, Khuzestan Agricultural Sciences and Natural Resources University, Mollasani, Khuzestan, Iran
phd.behdani@ramin.ac.ir
Mostafa
Ghaderi-Zefrehei
Department of Animal Science, Yasouj University, Yasouj, Iran
mghaderi@yu.ac.ir
Farjad
Rafeie
0000-0001-8083-0773
Department of Agricultural Biotechnology, Faculty of Agricultural Sciences, University of Guilan, Rasht, Iran
farjad.rafeie@gmail.com
Mohammad Reza
Bakhtiyarizadeh
Department of Animal and Poultry Science, College of Aburaihan, University of Tehran, Tehran, Iran
mrbakhtiari@ut.ac.ir
Hedayatallah
Roshanfekr
Department of Animal Science, Faculty of Animal and Food Science, Khuzestan Agricultural Sciences and Natural Resources University, Mollasani, Khuzestan, Iran
roshanfekr_hd@yahoo.com
Jamal
Fayazi
Department of Animal and Poultry Science, College of Aburaihan, University of Tehran, Tehran, Iran
j_fayazi@ramin.ac.ir
10.29252/ijb.1748
<strong>Background:</strong> The stress is one of main factors effects on production system. Several factors (both genetic and environmental elements) regulate immune response to stress.<br /> <strong>Objectives:</strong> In order to determine the major immune system regulatory genes underlying stress responses, a learning Bayesian network approach for those regulatory genes was applied to RNA-Seq data from a bovine leukocyte model system.<br /> <strong>Material and Methods:</strong> The transcriptome dataset GSE37447 was used from GEO and a Bayesian network on differentially expressed genes was learned to investigate the gene regulatory network.<br /> <strong>Results:</strong> Applying the method produced a strongly interconnected network with four genes (<em>TERF2IP, PDCD10, DDX10 </em>and<em> CENPE</em>) acting as nodes, suggesting these genes may be important in the transcriptome regulation program of stress response. Of these genes <em>TERF2IP</em> has been shown previously to regulate gene expression, act as a regulator of the nuclear factor-kappa B (NF-κB) signalling, and to activate expression of NF-κB target genes; <em>PDCD10</em> encodes a conserved protein associated with cell apoptosis; <em>DDX10</em> encodes a DEAD box protein and is believed to be associated with cellular growth and division; and <em>CENPE</em> involves unstable spindle microtubule capture at kinetochores. Together these genes are involved in DNA damage of apoptosis, RNA splicing, DNA repairing, and regulating cell division in the bovine genome. The topology of the learned Bayesian gene network indicated that the genes had a minimal interrelationship with each other. This type of structure, using the publically available computational tool, was also observed on human orthologous genes of the differentially expressed genes.<br /> <strong>Conclusions:</strong> Overall, the results might be used in transcriptomic-assisted selection and design of new drug targets to treat stress-related problems in bovines.
Cattle,genes,RNA,Stress
https://www.ijbiotech.com/article_85175.html
https://www.ijbiotech.com/article_85175_dd0eed6999f38ff8931d24399f1155d5.pdf
National Institute of Genetic Engineering and Biotechnology of Iran
Iranian Journal of Biotechnology
1728-3043
2322-2921
17
3
2019
09
01
Biodecolorization and Biodegradation of Azo Dye Reactive Orange-16 by Marine Nocardiopsis sp.
18
26
EN
Vaibhavi
Chittal
Biomolecules Laboratory, Technology Tower (TT 635), Vellore Institute of Technology (VIT) University, Vellore- 632014, Tamil Nadu, India
vaibhavichittal139@gmail.com
Magaly
Gracias
Biomolecules Laboratory, Technology Tower (TT 635), Vellore Institute of Technology (VIT) University, Vellore- 632014, Tamil Nadu, India
magalygracias21@gmail.com
Anagha
Anu
Biomolecules Laboratory, Technology Tower (TT 635), Vellore Institute of Technology (VIT) University, Vellore- 632014, Tamil Nadu, India
Purbasha
Saha
Biomolecules Laboratory, Technology Tower (TT 635), Vellore Institute of Technology (VIT) University, Vellore- 632014, Tamil Nadu, India
purbasha.saha@vit.ac.in
K.V.
Bhaskara Rao
Biomolecules Laboratory, Technology Tower (TT 635), Vellore Institute of Technology (VIT) University, Vellore- 632014, Tamil Nadu, India
10.29252/ijb.1551
<strong>Background:</strong> Azo dyes are xenobiotic compounds that have bioaccumulated in the environment due to escalated industrial development. These are hazardous in nature, possessing carcinogenic and mutagenic effects on human beings.<br /> <strong>Objectives:</strong> The perspective of the present study was to isolate and to determine azo dye (Reactive Orange-16) degrading potential of marine actinobacteria isolated from sediment samples of Port Blair, India.<br /> <strong>Material and Methods:</strong> Actinobacteria with dye decolorization potential were isolated from sea sediment samples. The actinobacterial isolate with the highest dye decolorizing percentage was identified with the help of phenotypic, biochemical and molecular studies. The different physico-chemical parameters for dye decolorization were also optimized. The nature of decolorization by the potent isolate was determined with the help of High Performance Liquid chromatography (HPLC) and Fourier Transformed Infrared spectroscopy (FTIR) techniques. Further the toxicity of RO-16 decolorized products was investigated with the help of phytotoxcity assay.<br /> <strong>Results:</strong> Out of six actinobacterial isolates, VITVAMB 1 possessed the most efficient RO-16 decolorization property. It decolorized 85.6% of RO-16 (250 mg L<sup>-1</sup>) within 24hrs. Isolate VITVAMB 1 was identified to be <em>Nocardiopsis </em>sp. Maximum dye decolorization occurred at pH 8, temperature 35°C, 3% salt concentration and a dye concentration of 50 mg L<sup>-1</sup>.<br /> <strong>Conclusions:</strong> The nature of decolorization by <em>Nocardiopsis</em> sp. was biodegradation. Additionally, the degraded dye metabolites were found to be less toxic than pure dye. The high decolorization potential of VITVAMB 1 and the low toxicity of its degradation products make it a prospective dye removal system. The marine origin of VITVAMB 1 also makes it an attractive source for novel azo dye reducing enzymes.
Biodegradation, Environmental,Spectroscopy, Fourier Transform Infrared,Chromatography, High Pressure Liquid,Reactive Orange-16
https://www.ijbiotech.com/article_95369.html
https://www.ijbiotech.com/article_95369_fadcdbfb1ebd36295c84008d2067556c.pdf
National Institute of Genetic Engineering and Biotechnology of Iran
Iranian Journal of Biotechnology
1728-3043
2322-2921
17
3
2019
09
01
Comparative Proteomic Analysis of Two Manilkara Species Leaves Under NaCl Stress
27
34
EN
Yumei
Liu
Xiamen overseas Chinese subtropical plant introduction garden, Xiamen, Fujian, China
xcong@163.com
Chongling
Yan
Key Laboratory of the Ministry of Education for Coastal and Wetland Ecosystems, Xiamen University, Xiamen, Fujian, China
ycl@xmu.edu.cn
Zhiyu
Song
Fujian Institute of Subtropical Botany, Xiamen, Fujian, China
songdiyang@163.com
Shuang
Zhou
Fujian Institute of Subtropical Botany, Xiamen, Fujian, China
angelazhou@xmu.edu.cn
10.29252/ijb.2219
<strong>Background:</strong> Salinity is a major environmental limiting factor, which affect agricultural production. The two <em>Manilkara </em>seedlings (<em>M. roxburghiana</em> and <em>M. zapota</em>) with high economic importance, could not adapt well to higher soil salinity and little is known about their proteomic mechanisms.<br /> <strong>Objectives: </strong>The mechanisms responsible for the effects of salinity on the two <em>Manilkara </em>species leaves were examined by means of proteomic analysis.<br /> <strong>Material and Methods:</strong> The seedlings were cultivated in a greenhouse and treated with NaCl. Leaves of control and the salt-stressed seedlings were sampled for phenol protein extraction. Proteins were separated by two-dimensional gel electrophoresis coupled with mass spectroscopy to study the change of proteins under different NaCl concentration.<br /> <strong>Results:</strong> For <em>M. roxburghiana</em> leaves, 21 protein spots exhibited significant abundance variations between the control and the 6‰, 8‰ NaCl treatments, of these 13 proteins were identified. They included L-ascorbate peroxidase, chloroplast carbonic anhydrase, phosphoglycerate kinase, 5 heat-shock proteins(HSPs) which were all down-regulated; For <em>M. zapota</em> leaves, 35 protein spots exhibited significant abundance variations, then 24 proteins were identified, including 7 down-regulated HSPs as well as glyceraldehyde-3-phosphate dehydrogenase, Cell division protein, putative mitochondrial NAD-dependent malate dehydrogenase, ATP synthase, Rubisco large subunit-binding protein, Cytochrome c peroxidase.<br /> <strong>Conclusions:</strong> Based on the common identified proteins between the two <em>M. </em>species, our results indicated that the identificated proteins in the two <em>Manilkara </em>species were involved in carbohydrate metabolism, photosynthesis, defense and stress. HSPs exhibited variation strictly related to NaCl stress. The down-regulated HSPs meant the function to repair cells that have suffered damage weaken during stress process. Furthermore, except for HSP70 in<em> M. zapota</em> leaves, the HSPs in the two species were all small heat shock proteins (sHSPs) with molecular weights ranging from 15 to 42 kDa.
2-DE,MS,Manilkara roxburghiana,Manilkara zapota
https://www.ijbiotech.com/article_91764.html
https://www.ijbiotech.com/article_91764_3f65fed02ca4b5362f3c4d207f9dc0e2.pdf
National Institute of Genetic Engineering and Biotechnology of Iran
Iranian Journal of Biotechnology
1728-3043
2322-2921
17
3
2019
09
01
Evaluation of Immunogenicity, Protective Immunity on Aquaculture Pathogenic Vibrio and Fermentation of Vibrio alginolyticus Flagellin FlaC Protein
35
42
EN
Chen
Chen
Chinese-German joint Institute for natural product research / Shaanxi Engineering Research Center for Tall Gastrodia Tuber and Medical Dogwood / College of Biological Science and Engineering, Shaanxi University of Technology, Hanzhong, China
Chao
Kang
Chinese-German joint Institute for natural product research / Shaanxi Engineering Research Center for Tall Gastrodia Tuber and Medical Dogwood / College of Biological Science and Engineering, Shaanxi University of Technology, Hanzhong, China
Na
Rong
Chinese-German joint Institute for natural product research / Shaanxi Engineering Research Center for Tall Gastrodia Tuber and Medical Dogwood / College of Biological Science and Engineering, Shaanxi University of Technology, Hanzhong, China
Nana
Wu
Chinese-German joint Institute for natural product research / Shaanxi Engineering Research Center for Tall Gastrodia Tuber and Medical Dogwood / College of Biological Science and Engineering, Shaanxi University of Technology, Hanzhong, China
Chunlin
Chen
Chinese-German joint Institute for natural product research / Shaanxi Engineering Research Center for Tall Gastrodia Tuber and Medical Dogwood / College of Biological Science and Engineering, Shaanxi University of Technology, Hanzhong, China
Sanqiao
Wu
Chinese-German joint Institute for natural product research / Shaanxi Engineering Research Center for Tall Gastrodia Tuber and Medical Dogwood / College of Biological Science and Engineering, Shaanxi University of Technology, Hanzhong, China
Xiaoying
Zhang
Chinese-German joint Institute for natural product research / Shaanxi Engineering Research Center for Tall Gastrodia Tuber and Medical Dogwood / College of Biological Science and Engineering, Shaanxi University of Technology, Hanzhong, China
zhang.xy@nwsuaf.edu.cn
Xiang
Liu
Chinese-German joint Institute for natural product research / Shaanxi Engineering Research Center for Tall Gastrodia Tuber and Medical Dogwood / College of Biological Science and Engineering, Shaanxi University of Technology, Hanzhong, China
zhang@bio.uminho.pt
10.29252/ijb.2628
<strong>Background:</strong> <em>Vibrio </em>are the main pathogenic bacteria in aquaculture. The flagellin protein C (FlaC) of <em>Vibrio alginolyticus</em> has good immunogenicity and the prospect of potential application in a vaccine.<br /> <strong>Objectives: </strong>We aimed to evaluate the immunogenicity, protective immunity, and prokaryotic expression fermentation of <em>V. alginolyticus</em> FlaC protein for the vaccine in aquaculture.<br /> <strong>Material and Methods:</strong> A molecular cloning method was used to construct the expression strain of FlaC protein, and the protein was purified with Ni-affinity chromatography. Polyclonal antiserum was prepared via mice immunized with the FlaC protein. The Western blot and enzyme-linked immunosorbent assay (ELISA) were used to check the specificity and titre of the antiserum. ELISA and pull-down assay detected the interaction between FlaC protein antiserum and <em>Vibrio</em>. The immune protection function of FlaC protein was detected with mice actively immunized with FlaC protein and challenged by <em>V. alginolyticus </em>and<em> V. parahaemolyticus</em>. The optimal expression conditions for FlaC protein were detected using an L<sub>9</sub>(3<sup>4</sup>) orthogonal design model.<br /> <strong>Results:</strong> The expression strain of FlaC protein was obtained successfully, and purified FlaC protein was prepared using a mice polyclonal antibody. The FlaC protein antiserum held a high specificity, and the titre was 1:3200. The antiserum directly interacted with <em>V. alginolyticus</em> and <em>V. parahaemolyticus</em>, and the FlaC protein demonstrated a significant immune protection function (50%) against <em>V. alginolyticus</em> infection and some immune protection function (41.66%) against <em>V. parahaemolyticus</em>. The optimal expression conditions for FlaC protein included a strain OD<sub>600</sub> value of 0.8, final isopropyl-<em>β</em>-d-thiogalactoside (IPTG) concentration of 0.1 mmol/L, an inducing time of 8 hours, and an inducing temperature of 28°C.<br /> <strong>Conclusions:</strong> This study showed that the FlaC protein possesses a significant immunogenicity and immune protection effect and obtained the optimal fermentation conditions. It is expected to be a potential vaccine against <em>V. alginolyticus</em> and <em>V. parahaemolyticus</em>.
Vibrio alginolyticus,FlaJ Protein,Vibrio parahaemolyticus
https://www.ijbiotech.com/article_95370.html
https://www.ijbiotech.com/article_95370_e6974c2cae6c54edf4790a6245c2f716.pdf
National Institute of Genetic Engineering and Biotechnology of Iran
Iranian Journal of Biotechnology
1728-3043
2322-2921
17
3
2019
09
01
Multi-function Plant Defensin, Antimicrobial and Heavy Metal Adsorbent Peptide
43
49
EN
Neda
Mirakhorli
Department of Plant Breeding and Biotechnology, Shahrekord University, Shahrekord, Iran
mirakhorli-n@agr.sku.ac.ir
Zahra
Norolah
Department of Plant Breeding and Biotechnology, Shahrekord University, Shahrekord, Iran
z.nurolah@gmail.com
Samira
Foruzandeh
Department of Plant Breeding and Biotechnology, Shahrekord University, Shahrekord, Iran
samiraforuzandeh65@yahoo.com
Fateme
Shafizade
Department of Plant Breeding and Biotechnology, Shahrekord University, Shahrekord, Iran
asemaneshargi@gmail.com
Farzaneh
Nikookhah
Department of Fisheries and Environmental Science, Shahrekord University, Shahrekord, Iran
f_nikookhah@yahoo.com
Behnaz
Saffar
Department of Genetic, Shahrekord University, Shahrekord, Iran
saffar_b@sci.sku.ac.ir
Omid
Ansari
Ecofibre Industries Operations and Ananda Hemp, Brisbane, Australia
omid@ecofiber.com
10.29252/ijb.1562
<strong>Background:</strong> Defensin peptide isolated from plants are often heterogeneous in length, sequence and structure, but they are mostly small, cationic and amphipathic. Plant defensins exhibit broad-spectrum antibacterial and antifungal activities against Gram-positive and Gram-negative bacteria, fungi and etc. Plant defensins also play an important role in innate immunity, such as heavy metal and some abiotic stresses tolerance.<br /> <strong>Objectives:</strong> In this paper, <em>in vitro</em> broad-spectrum activities, antimicrobial and heavy metal absorption, of a recombinant plant defensin were studied.<br /> <strong>Material and Methods:</strong> <em>SDmod</em> gene, a modified plant defensin gene, was cloned in pBISN1-IN (EU886197) plasmid, recombinant protein was produced by transient expression via Agroinfiltration method in common bean. The recombinant protein was tested for antibacterial activity against Gram-negative, Gram-positive bacteria and <em>Fusarium</em> sp. the effects of different treatments on heavy metal zinc absorption by this peptide were tested.<br /> <strong>Results: </strong>We confirmed the antibacterial activities of this peptide against Gram-negative (<em>Escherichia</em> <em>coli</em> and <em>Pseudomonas aeruginosa</em>) and Gram-positive (<em>Staphylococcus aureus </em>and <em>Bacillus cereus</em>) bacteria, and antifungal activities of this peptide against <em>Fusarium</em> spp. (<em>Fusarium oxysporu</em><em>m</em> and <em>Fusarium solani).</em> High metal absorption coefficient for this peptide was also observed.<br /> <strong>Results:</strong> Out of six actinobacterial isolates, VITVAMB 1 possessed the most efficient RO-16 decolorization property. It decolorized 85.6% of RO-16 (250 mg L<sup>-1</sup>) within 24hrs. Isolate VITVAMB 1 was identified to be <em>Nocardiopsis </em>sp. Maximum dye decolorization occurred at pH 8, temperature 35<sup>o</sup>C, 3% salt concentration and a dye concentration of 50 mg L<sup>-1</sup>.<br /> <strong>Conclusions:</strong> Results suggesting that modified defensin peptide facilitates a broader range of defense activities. dedefensins are an important part of the innate immune system in eukaryotes. These molecules have multidimensional properties that making them promising agents for therapeutic drugs.
Antibacterial Effect,Heavy metal adsorption,Plant defensin,Therapeutic potential
https://www.ijbiotech.com/article_91759.html
https://www.ijbiotech.com/article_91759_6cb218806489582b9d0b2a19a475f1be.pdf
National Institute of Genetic Engineering and Biotechnology of Iran
Iranian Journal of Biotechnology
1728-3043
2322-2921
17
3
2019
09
01
Gene-Based Marker to Differentiate Among B, A, and R Lines in Hybrid Production of Rapeseed Ogura System
50
54
EN
Javad
Akbari Afjani
Department of Biotechnology, Faculty of Agriculture, Tarbiat Modarres University, Tehran, Iran
javadakbari89@yahoo.com
Masood
Soltani Najafabadi
0000-0002-9634-9981
Seed and Plant Improvement Institute (SPII), Agricultural Research, Education and Extension (AREEO), Shahid Fahmideh BLVRD, Seed and Plant Improvement Institute Campus, Karaj, Iran
m.soltani@spii.ir
Reza
Gholi Mirfakhraei
Department of Plant Genetics and Breeding, Faculty of Agriculture, Tarbiat Modarres University, Tehran, Iran
abdhoorz@modares.ac.ir
10.29252/ijb.1870
<strong>Background:</strong> In plant breeding program to produce hybrid varieties, pair of male sterile and restorer fertility lines are required. Differentiation of lines possessing restorer fertility allele from the lines lacking it remove the need for the progeny test, and thus reducing the time and the cost in the hybrid production procedure. Canola breeding program in Iran has concentrated toward production of domestic hybrid varieties, however, it suffers from lack of molecular information in restore fertility status of lines, and therefore it needs time and tedious activities.<br /> <strong>Objectives:</strong> To design gene-based markers for distinguishing R-, A- lines and hybrids in sunflower breeding programs.<br /> <strong>Material and Methods:</strong> Aligning sequences of locus responsible for male sterility and that of male fertility resulted in finding differences in the loci, which used to define two set of suitable primer pairs. Genomic DNA from 25 R-lines (23 inbred lines and two commercial lines), 9 A-lines (7 inbred lines and two commercial lines), one B-line and two commercial hybrids were extracted and used in PCR as template.<br /> <strong>Results:</strong> Using one-primer pairs, a band of nearly 1500 bp was amplified in restorer lines but not in A-, B- lines. Another primer pair used to distinguish hybrids (heterozygout) from restorer lines. Results of the report is predicted to be used in canola breeding for hybrid production.<br /> <strong>Conclusions:</strong> Although the molecular bases for the male sterility and fertility restoration in rapeseed is not published, taking advantages of gene-based markers, make rapeseed breeding program more efficient regarding time and costs.
Biomarkers,Brassica rapa, Rf Allele
https://www.ijbiotech.com/article_85172.html
https://www.ijbiotech.com/article_85172_55da68799eb9a32c98d0de48aeabe185.pdf
National Institute of Genetic Engineering and Biotechnology of Iran
Iranian Journal of Biotechnology
1728-3043
2322-2921
17
3
2019
09
01
Role of Molecular Interactions and Oligomerization in Chaperone Activity of Recombinant Acr from Mycobacterium tuberculosis
55
62
EN
Gautam
Krishnan
0000-0002-1064-3056
Department of Biological Sciences, BITS Pilani KK Birla Goa Campus, Zuari Nagar, Goa 403726, Goa, India
krishnangautam@gmail.com
Utpal
Roy
Department of Biological Sciences, BITS Pilani KK Birla Goa Campus, Zuari Nagar, Goa 403726, Goa, India
utpalroy010@gmail.com
10.29252/ijb.2370
<strong>Background:</strong> The chaperone activity of <em>Mycobacterium tuberculosis </em>Acr is an important function that helps to prevent misfolding of protein substrates inside the host, especially in conditions of hypoxia.<br /> <strong>Objectives: </strong>The aim of this study was to establish the correlation of structure and function of recombinant Acr proteins both before and after gel filtration chromatography. The aim was also to find the oligomeric conformation of these samples and use this information to explain differences in activity<br /> <strong>Material and Methods:</strong> <em>M. tuberculosis acr</em> gene was cloned with an N-terminal His-tag in pET28a and expressed with IPTG induction in BL2 (DE3) competent<em> Escherichia coli</em>. The activity of a recombinant Acr without gel filtration was checked by preventing thermal aggregation of citrate synthase at 45°C and the chaperone activity against insulin B chain aggregation at 60°C and 37°C. On further purification using gel filtration chromatography, the protein was again tested for chaperone activity using insulin as substrate at 37°C with two types of samples without and with gel filtration designated A and B respectively. The effects of pre–heat treatment at 60 °C on chaperone activity of both A and B samples were studied by performing the chaperone assay at 37°C.<br /> <strong>Results:</strong> The level of expression was 40 to 50 mg /l. The protein was expressed in a soluble form at 37°C and subsequently purified by a 3 step gradient of imidazole using Ni-NTA resin. Gel filtration chromatography showed recombinant Acr to be a mixture of 9 to 15-mers, whereas Native-PAGE analysis showed a large proportion of 5 and 7 mers in the non gel-filtered sample, while non gel –filtered samples showed more proportions of higher size oligomers. The chaperone activity of non gel-filtered (A) samples was less than gel-filtered (B) samples at 37°C with 24 µM required of A for complete inhibition as compared to 6 µM of B. The chaperone activity of non gel–filtered samples at 60°C showed complete inhibition of activity at a concentration of 44 µM. Molecular interaction studies showed influence of size of oligomers on molecular coverage of insulin B chain. Pre-heat treatment improved the activity only after the gel filtration.<br /> <strong>Conclusions:</strong> The larger proportion of monomers in the non gel-filtered sample could explain the difference in activity as compared to the gel-filtered samples in terms of molecular interaction with insulin. Increased oligomer size favorably affected secondary structure, a finding not reported so far, and warranting further investigation. A molecular level interaction of inhibition was predicted using Avogadro number of molecules and oligomer size. The difference in activity after pre–heat treatment seemed to indicate an important role for oligomerization.
Acr,insulin,chaperone,oligomer,pre-heat treatment,Mycobacterium Tuberculosis
https://www.ijbiotech.com/article_85200.html
https://www.ijbiotech.com/article_85200_651f63f41f0ba2a0659aa3f8fadff03d.pdf
National Institute of Genetic Engineering and Biotechnology of Iran
Iranian Journal of Biotechnology
1728-3043
2322-2921
17
3
2019
09
01
New Brucella abortus S19 Mutant to Improve Distinction Between Infected and Vaccinated Animals
63
67
EN
Bahar
Nayeri Fasaei
0000-0003-2373-4667
Department of Microbiology, Faculty of Veterinary Medicine, University of Tehran, Tehran, Iran
nayerib@ut.ac.ir
Soulmaz
Naserli
Department of Microbiology, Faculty of Veterinary Medicine, University of Tehran, Tehran, Iran
soulmaz.naserli@yahoo.com
Taghi
Zahrai Salehi
0000-0002-5665-5757
Department of Microbiology, Faculty of Veterinary Medicine, University of Tehran, Tehran, Iran
tsalehi@ut.ac.ir
Alireza
Saeedinia
Department of Bioscience and Biotechnology, Malek-Ashtar University of Technology, Tehran, Iran
a.saeedinia@modares.ac.ir
Alimohammad
Behroozikhah
Department of Brucellosis, Razi Vaccine and Serum Research Institute, Agricultural Research Education and Extension Organization (AREEO), Karaj, Iran
2112behroozikhah@gmail.com
Iradj
Ashrafi Tamai
Department of Microbiology, Faculty of Veterinary Medicine, University of Tehran, Tehran, Iran
iradjashrafi@gmail.com
10.29252/ijb.2159
<strong>Background:</strong> Using <em>Brucella abortus</em> Strain 19 (S19) to control bovine brucellosis is restricted due to induce antibodies to the O-side chain of the smooth lipopolysaccharide (LPS) which may be difficult to differentiate vaccinated and infected animals. Furthermore, it is virulent for humans and can induce abortion to cattle.<br /> <strong>Objectives:</strong> The aim of this study was to employ gene knockout <em>B. abortus</em> S19 for the first time to eliminate diagnostic defects and obtain the attenuated mutant strain.<br /> <strong>Material and Methods:</strong> The <em>wbkA</em> gene, which is one of the LPS O-chain coding genes, was knocked out in vaccinal <em>Brucella abortus</em> S19. The proliferative response and immunoglobulin M production were analyzed in<em> wbkA</em> deletion strain-infected BALB/c mice.<br /> <strong>Results:</strong> The loss of <em>wbkA</em> gene function resulted in induction of the splenocyte proliferative response in mice infected by the mutant S19 strain compare to those induced by parental S19 and RB51 strains. Moreover, <em>wbkA</em> mutant did not induce any IgM antibody response using the enzyme-linked immunosorbent assay.<br /> <strong>Conclusions:</strong> As a result, the new mutant S19 strain had deficiency in its LPS O-chain structure, besides cannot induce IgM response then, reduce mistakes to discriminate between vaccinated and infected animal, and also can be considered as a new vaccine candidate.
Brucella abortus,Mutant Strains,Gene Knockout Techniques
https://www.ijbiotech.com/article_91762.html
https://www.ijbiotech.com/article_91762_42d42c948c3d66b5c17b5c0fc418c8ca.pdf
National Institute of Genetic Engineering and Biotechnology of Iran
Iranian Journal of Biotechnology
1728-3043
2322-2921
17
3
2019
09
01
Evaluation of ten SNP Markers for Human Identification and Paternity Analysis in Persian Population
68
71
EN
Sajad
Habibi
Human Genetic Research Center, Baqiatallah University of Medical Sciences, Tehran, Iran
s.habibi28@yahoo.com
Amirhossein
Ahmadi
Department of Genetics, Faculty of Sciences, Persian Gulf University, Bushehr, Iran
a_ahmadi1985@yahoo.com
Mehrdad
Behmanesh
0000-0002-3901-304X
Department of Genetics, Faculty of Biological Sciences, Tarbiat Modares University, Tehran, Iran
behmanesh@modares.ac.ir
Ali
Miri
Human Genetic Research Center, Baqiatallah University of Medical Sciences, Tehran, Iran
alimiri1391@gmail.com
Mahmoud
Tavallaie
0000-0003-0080-4298
Human Genetic Research Center, Baqiatallah University of Medical Sciences, Tehran, Iran
tavalla.mah@gmail.com
10.29252/ijb.2148
<strong>Background:</strong> DNA markers are inevitable tools of human identification in forensic science. Single Nucleotide Polymorphisms (SNPs) are one category of these markers which is concerned to use especially in the case of degraded DNA because of their short amplicons.<br /> <strong>Objectives:</strong> Detection of highly informative SNPs by the criteria is the essential step to develop a useful panel of SNP markers. The purpose of this work is to get high informative SNPs for human identification in Persian ethnic of the Iranian population.<br /> <strong>Material and Methods:</strong> Genotype and allele frequencies of 10 SNPs from the SNPforID browser were determined by a PCR-RFLP method on 100 samples that was taken from 100 unrelated Persian people.<br /> <strong>Results:</strong> These ten SNPs were in Hardy-Weinberg equilibrium (<em>P </em>value > 0.1) except rs1355366 (<em>P </em>value = 0.02) and Heterozygosity of seven SNPs is greater than 0.45 but minor allele frequency of only four SNPs is more than 0.45. According to criteria only three SNPs rs1454361, rs2111980 and rs2107612 can pass all standards and are highly informative in population for forensic uses.<br /> <strong>Conclusions:</strong> Our data showed that the CPI (Combined probability of Identity) and CPE (Combined Power of Exclusion) for ten SNPs are 1.13 E-04 and 0.809 respectively. It was also showed based on the criteria only three SNPs (rs2107612, rs1454361 and rs2111980) are highly informative in Persian population. If we can find 39 SNPs with PE and PI close to PE and PI of these three SNPs (rs2107612, rs1454361 and rs2111980), we will be able to use of these 39 SNPs in human identification with sufficient power of discrimination.
Forensic Sciences,Forensic Anthropology,Polymorphism, Single Nucleotide
https://www.ijbiotech.com/article_91763.html
https://www.ijbiotech.com/article_91763_f6e5d341642777a3e3be8ae755cad075.pdf