2024-03-29T02:46:54Z
https://www.ijbiotech.com/?_action=export&rf=summon&issue=1158
Iranian Journal of Biotechnology
IRAN J BIOTECH
1728-3043
1728-3043
2005
3
1
A New Reporter Gene Technology: Opportunities and Perspectives
Gholam Reza
Salehi Jouzani
Irina
Vasilievna Goldenkova
The paper summarizes the current status of the reporter gene technology and their basics. Reporter gene technology is widely used to monitor cellular events associated with gene expression and signal transduction. Based upon the splicing of transcriptional control elements to a variety of reporter genes, it “reports” the effects of a cascade of signaling events on gene expression inside cells. The principal advantage of these assays is their high sensitivity, reliability, convenience and adaptability to large scale measurements.The reporter gene systems β-galactosidase (LacZ), luciferases (LUC), β-glucoronidase (GUS),green fluorescent protein (GFP) and GFP-like proteins, widely used presently, are compared. The potentialitiesof a new reporter system, based on thermostable lichenase (β-1, 3-1, 4-gluconases) of Clostridium thermocellum are described in detail. Advantages and disadvantages of the used reporter systems, and also the opportunities for development and application of bifunctional reporter systems are considered. Perspectives of reporter genes for studying gene expression regulation in prokaryotic and eukaryotic organisms are elucidated.
Reporter genes
β-galactosidase (LacZ)
Luciferase (LUC)
β-glucoronidase (GUS)
Green fluorescent protein (GFP)
GFP-like proteins, Lichenase (LicB)
2005
01
01
1
15
https://www.ijbiotech.com/article_6961_1c3955af5df42f923dedbc2943e294d5.pdf
Iranian Journal of Biotechnology
IRAN J BIOTECH
1728-3043
1728-3043
2005
3
1
Bacterial Expression and Functional Characterization of A Naturally Occurring Exon6-less Preprochymosin cDNA
Gholamreza
Ahmadian
Mehdi
Arbabi Ghahroudi
Nasrin
Rastgoo
Ghodrat
Rahimi Mianji
Chymosin (Rennin EC 3.4.23.4), an aspartyl proteinase, is the major proteolytic enzyme in the fourthstomach of the unweaned calf, and it is formed by proteolytic activation of its zymogene, prochymosin.Following the cloning of synthesized cDNAs on mRNA pools extracted from the mucosa of the calf fourthstomach, we have identified an alternatively spliced form of preprochymosin cDNA (AS6 preprochymosin).Sequencing data analysis showed that the exon six has been spliced out and, therefore the gene productis 114 bp shorter in length. In order to determine the biological significance of the AS6 preprochymosin, weexpressed the encoding cDNA together with a complete chymosin cDNA in E. coli. Under the same expression conditions, we found at least a 5-fold higher expression of AS6 preprochymosin protein in comparisonto a full-length recombinant chymosin. Protein prediction program analyses showed that the missingexon contain groups of amino acids with high hydrophobicity score. Therefore, the deletion of this exon may explain the higher expression of the recombinant product in E. coli. Most importantly, the biological activity of the purified AS6 preprochymosin, was confirmed in an assay of chymosin milk-clotting activity using the recombinant preprochymosin and commercial rennet as positive controls. The expression of the biologically active preprochymosin lacking exon 6 may have important implications on the existence of this splicing form of mRNA in vivo and on its biotechnological applications in cheese manufacture.
Preprochymosin
Aspartyl proteinase
Alternatively spliced transcript
Milk-clotting
Microbial rennet
Recombinant chymosin
Hydrophobicity
2005
01
01
16
23
https://www.ijbiotech.com/article_6946_a69223ed5dd30e915c09a0f13e170566.pdf
Iranian Journal of Biotechnology
IRAN J BIOTECH
1728-3043
1728-3043
2005
3
1
Evaluation of Heat Induction Strategy for Recombinant Human Growth Hormone Expression in Fed-Batch Fermentation
Fatemeh
Tabandeh
Seyed Abbas
Shojaosadati
Bagher
Yakhchali
Mahvash
Khodabandeh
Mohammad Hossein
Sanati
The high cell density cultivation (HCDC) of a recombinant E. coli producing human growth hormone (hGH)under different heat induction strategies has been reported. In this paper the effect of heat shock temperature, its duration and post-induction temperature, at two levels on cell growth and death, hGH production and its degradation, substrate utilization and by-product formation were investigated by full factorial experimental design. The results showed that heat shock temperature was the most effective factor on hGH production having approximately 75.8% of total contribution. There was no significant accumulation of substrate or by-product in culture medium during HCDC in all experiments. Thirty two percent of cells were subjected to lysis during the heat induction at 42°C for 20 min followed by 37°C for 4h. Biologically active recombinant hGH was produced comprising 13% of total cell protein without degradation. An empirical equationwas employed to describe the relationship between three factors and hGH production during HCDC.
Heat induction
High cell density culture
Human growth hormone
recombinant E. coli
2005
01
01
24
30
https://www.ijbiotech.com/article_6960_e4730112f658c84cec3f1ba8efc827b9.pdf
Iranian Journal of Biotechnology
IRAN J BIOTECH
1728-3043
1728-3043
2005
3
1
Partial Epitope Mapping of Alfalfa mosaic Virus and the Effect of Coat Protein Gene Mutation on Aphid Transmission
Hossain
Massumi
Phil
Jones
Nigle
Hague
Epitope mapping with seventy-one overlapping octapeptides representing the whole sequence of Alfalfa mosaic virus (AMV) coat protein (CP) (strain S) was done using monoclonal antibodies. In total, five monoclonal antibodies identified 5 epitopes, at different sites along the amino acid sequence of AMV coatprotein. Four MAbs each reacted with a single epitope, while one MAb bonded with peptides on twowidely separated parts of the coat protein. A full length DNA copy of RNA 3 of AMV strain S in a pBS plasmidwas used as a template for mutation and transcription. Both before and after modification, the RNA 3 of thisstrain was inoculated to Nicotiana tabacum transgenic P12 plants and AMV particles were purified. Strain Sof AMV produced systemic symptoms in N. tabacum transgenic P12 plants. It was suggested that MAb-2was effective in blocking insect transmission of AMV. Epitope-2 which play an important role in transmissionis recognized by MAb-2. To further investigate the interaction between epitope-2 and transmission ofAMV mutation of the amino acids of epitope-2 was made in the appropriate coding regions of the coatprotein of AMV strain S. These changes were (i) a phenylalanine to tyrosine and (ii) asparagine to glutamine.The mutation of asparagine to glutamine had no effect on the transmission of AMV by M. persicae,but the mutation of phenylalanine to tyrosine gave a significant reduction (P = 0.02) in aphid transmissibility.This indicated that the phenylalanine residue at position 67 of the AMV coat protein was involved in transmission.
Alfalfa mosaic virus
Coat protein
epitope mapping
Transmission
Mutation
2005
01
01
31
40
https://www.ijbiotech.com/article_6938_5fee9fd58cc4f71f2212ce6df57132a5.pdf
Iranian Journal of Biotechnology
IRAN J BIOTECH
1728-3043
1728-3043
2005
3
1
Survey of Efficiency of Six Microsatellite Loci in Iranian Indigenous Cattle and Buffalo Populations
Seyed-Ziyaeddin
Mirhoseinie
Seyed-Mohammad Farhad
Vahidie
Behzad
Gharehyazie
Genetic diversity of three native buffalo populations from Azarie, Mazandaranie and Khuzestanie and twoIranian cattle breeds namely Sistanie and Taleshie were estimated using six microsatellite markers. Thirtyindividuals were randomly selected from each population/ breed. Total genomic DNA was extracted by anoptimized phenol–chloroform extraction method. The extracted DNA was amplified through polymerasechain reaction (PCR). Of the six microsatellite loci used in this study, two loci (ETH10 and ETH 225) weremonomorphic within the three buffalo populations. Genetic distance between the populations was estimatedby Fst (two by two) method. Maximum genetic distance was observed between Khuzestanie andMazandaranie buffalo populations (55%); whereas the minimum genetic distance (31%) was observedbetween Khuzestanie and Azarie populations. Values of both polymorphic information content (PIC) and heterozygosity (observed and expected) were higher within two cattle breeds as compared to those estimatedfor three buffalo populations.
genetic diversity
Polymorphism
Microsatellite markers
buffalo
Cattle
2005
01
01
41
47
https://www.ijbiotech.com/article_6942_5ebb5cac5cc010e1629e4c176e990a08.pdf
Iranian Journal of Biotechnology
IRAN J BIOTECH
1728-3043
1728-3043
2005
3
1
PCR Detection of Plasmid Mediated TEM, SHV and AmpC β-Lactamases in Community and Nosocomial Urinary Isolates of Escherichia coli
Fereshteh
Eftekhar
Seyed Mehdi
Hosseini-Mazinani
Soheila
Ghandili
Minoo
Hamraz
Shahrzad
Zamani
Fifty clinical isolates of Escherichia coli from two groups of subjects (hospitalized and outpatients) werestudied for their susceptibility to twelve β-lactam antibiotics. All isolates were resistant to ampicillin, amoxicillin, oxacillin and cefradine. Carbenicillin resistance was found in all but one outpatient isolate. Resistance to cephalothin, cephalexin and cefazoline ranged from 76% to 96% among the test bacteria. On the other hand, the majority of the isolates were sensitive to cefoxitime, ceftizoxime and ceftriaxone (84-100%). β-lactamases from all bacteria were inhibited by clavulanic acid and none harbored extended spectrum β-lactamases as shown by the double disc diffusion method. Gene amplification by polymerase chain reaction (PCR) using specific primers for AmpC, TEM and SHV enzymes showed that 92% of all organisms carried blaTEM alone, or along with SHV or ampC genes. Presence of all three genes was shown in 8% of the hospitalized patients and 20% of the outpatients. Conjugative transfer of β-lactam resistance markers into susceptible host recipients occurred for 72% of the isolates in each group. Overall, the antibiotic susceptibility profile, the distribution of the β-lactamase gene type and the rate of conjugative transfer of the resistance markers were similar in both subject groups.
Escherichia coli
TEM
SHV
AmpC
β-lactamases and PCR Typing
2005
01
01
48
54
https://www.ijbiotech.com/article_6962_1dc95d8482175c2e63361f9ac23b0361.pdf
Iranian Journal of Biotechnology
IRAN J BIOTECH
1728-3043
1728-3043
2005
3
1
cDNA Microarray Data Normalization
Mohammad Mehdi
Naghizadeh
Ebrahim
Hajizadeh
Anoshirvan
Kazemnejad
Normalization process is a set of operations whereby systematic biases are removed from microarray data.Therefore researcher can attain acceptable results and have more logic comparisons. This process consideringthe bias constructions and the effects on microarray is recognizable and applicable. Examplesof this process are spatial correction, background correction, rescaling, dye effect and within slide normalization. Spot intensity standard deviation is decreased by that normalization processes, and confidence to the microarray data analysis outcomes is increased. This paper describes all of these methods. In addition it contains the examples of a real dual channel cDNA microarray experiment to illustrate normalization process.
Microarray
Normalization
cDNA
preprocessing
Background correction
Robust smoother
2005
01
01
55
63
https://www.ijbiotech.com/article_6947_88ae17c4072842b8239a0df12fd7ee2f.pdf
Iranian Journal of Biotechnology
IRAN J BIOTECH
1728-3043
1728-3043
2005
3
1
CpG Motif as an Adjuvant in Immunization of a Recombinant Plasmid Encoding Hepatitis C Virus Core Protein
Ali
Karami
Mohammad Nabi
Sarbolouki
Fatemeh
Khatami
Nasrin
Rastgoo
The immunogenicity and protective efficacy of DNA vaccines have been demonstrated in numerous animalmodels of infectious diseases. In order to increase the potency of DNA vaccines, in this study, conventionaladjuvants such as aluminium phosphates, dendrosome, CpG motif and mixture of aluminium phosphateand CpG motif have been tested. Female BALB/c mice were immunized with mixture of 10, 25 and 50 μg HCVcore pcDNA3. Each dose of recombinant pcDNA3 together with different adjuvants used as an immunogenwere injected three times on day; 0, 30 and 50 days. Blood samples were collected at four different times intervals and antibody response against HCV core antigen was determined by HCV core ELISA kit. The results indicate that the best antibody response was with mixture of aluminium phosphate and CpG motif as an adjuvant. This data suggest that the antibody response induced following DNA immunization can be modified by formulation strategies.
CpG Motif
Dendrosome, Hepatitis C Virus
HCV Core pcDNA3
2005
01
01
64
66
https://www.ijbiotech.com/article_6937_83cb3be0e1734bdb3683703fdbc8fadc.pdf