eng
National Institute of Genetic Engineering and Biotechnology of Iran
Iranian Journal of Biotechnology
1728-3043
2322-2921
2003-01-01
1
1
1
18
6895
Mitochondrial DNA Mutations, Pathogenicity and Inheritance
Massoud Houshmand
housh62@yahoo.com
1
National Research Center for Genetic Engineering and Biotechnology,
Mitochondria contain their own DNA (mtDNA), which codes for 13 proteins (all subunits of the respiratory chain complexes), 22 tRNAs and 2 rRNAs. Several mtDNA point mutations as well as deletions have been shown to be causative in well-defined mitochondrial disorders. A mixture of mutated and wild type mtDNA (heteroplasmy) is found in most of these disorders. Inheritance of mtDNA is maternal, and mothers with heteroplasmic mtDNA transmit different proportions of normal and mutated mtDNA to the children.Mitochondrial tRNA genes have a central role in mitochondrial gene expression at the level of transcription,RNA processing and protein synthesis and they appear to be the mitochondrial genes most frequentlyaffected by mutations causing diseases in man.
https://www.ijbiotech.com/article_6895_78bec9803215c35e2b7467bc94498bda.pdf
mitochondria
MtDNA
Inheritance and Pathogenecity
eng
National Institute of Genetic Engineering and Biotechnology of Iran
Iranian Journal of Biotechnology
1728-3043
2322-2921
2003-01-01
1
1
19
25
6894
Diagnosis and Disease Management in CML Patients Using Conventional and Molecular Cytogenetics
Kiran Kucheria
1
Rashmi Talwar
2
Division of Genetics, Department of Anatomy, All India Institute of Medical Sciences,New Delhi
Division of Genetics, Department of Anatomy, All India Institute of Medical Sciences, New Delhi
Chronic Myeloid Leukemia (CML) is a hematopoietic malignancy characterized by the presence ofPhiladelphia (Ph1) chromosome that results from balanced reciprocal translocation between chromosomes9 and 22 leading in the formation of bcr/abl fusion gene. The present study was conducted to evaluate cytogenetic and molecular abnormalities in CML patients at presentation and during the course of therapy. Cytogenetic analysis was carried out in bone marrow samples of 165 suspected patients of CML using standard protocols. Sequential cytogenetic analysis was also done in 55 CML patients (50 on IFN-α 2b therapy and 5 on Hydroxyurea) up to variable period of 3 years. Fluorescence In Situ Hybridization (FISH) using specific probes for bcr and abl genes was carried out in cases where conventional cytogenetics was not informative, in cases that were Ph-negative at presentation and in cases with complete or major cytogenetic response (<34%Ph+cells). Of the 165 CML patients, 157 patients (95%) were Ph-positive while 8 patients (5%) werePh-negative. Cytogenetic abnormalities other than the standard Ph1 translocation were observed in 2Ph+ patients and 2 Ph-negative patients. Sequential cytogenetic analysis revealed varied degrees of cytogenetic response in 35 patients on IFN-α 2b therapy.Complete cytogenetic response was observed in 8 patients (16%) on IFN-α 2b therapy. However, FISHanalysis could detect bcr/abl fusion gene in some of these cases. This finding highlights the sensitivity ofthis technique in detecting residual disease even in patients with complete cytogenetic remission. Theother patients in complete cytogenetic remission did not have evidence of bcr/abl fusion. FISH analysisalso revealed bcr/abl fusion signals in Ph-negative cases (with no cytogenetic abnormality) indicating amasked translocation or a sub-microscopic rearrangement.Thus, the results of the present study show that analysis of cancer patients on therapy at the molecularlevel has tremendous importance for management of CML patients. Further, with the use of highly sensitive FISH technique, results can be obtained within 24 hours thereby, aiding rapid diagnosis and management of these patients. This efficient and highly sensitive molecular cytogenetic technique can also be done on interphase cells and poorly spread metaphases thereby, overcoming the difficulty of conventional chromosomal analysis especially in patients on therapy.
https://www.ijbiotech.com/article_6894_b62cdbfe978699328fa9066d15b5f780.pdf
CML
Diagnosis
cytogenetics
fish
eng
National Institute of Genetic Engineering and Biotechnology of Iran
Iranian Journal of Biotechnology
1728-3043
2322-2921
2003-01-01
1
1
26
30
6875
DHPLC Applications: Finding DNA Variation on the Y Chromosome
Qasim Ayub
1
Aisha Mohyuddin
2
Shagufta Khaliq
3
Abdul Hameed
ahz1963@yahoo.com
4
Chris Tyler Smith
5
S. Qasim Mehdi
6
Biomedical and Genetic Engineering Division, Dr. A. Q. Khan Research Laboratories, Islamabad
Biomedical and Genetic Engineering Division, Dr. A. Q. Khan Research Laboratories,
1Biomedical and Genetic Engineering Division, Dr. A. Q. Khan Research Laboratories, Islamabad
Biomedical and Genetic Engineering Division, Dr. A. Q. Khan Research Laboratories, Islamabad
CRC Chromosome Molecular Biology Group, Department of Biochemistry, University of Oxford, Oxford, UK.
Biomedical and Genetic Engineering Division, Dr. A. Q. Khan Research Laboratories, P.O. Box 2891, Islamabad 44000, Pakistan.
Denaturing High-Performance Liquid Chromatography (DHPLC) is a recently developed technique forthe detection of single nucleotide polymorphisms (SNPs) and mutations. It involves the comparisonbetween two or more DNAs as a mixture of denatured and reannealed PCR products. The methodologyis based on the principle of reversed phase liquid chromatography and uses a unique DNA separationmatrix. The exquisite sensitivity of the technique is determined by adjusting the oven temperature.Elution of DNA fragments is dependant on the chain length and sequence and can be predicted by computation.Under partially denaturing conditions heteroduplices formed upon mixing, denaturing, and reannealing of two, or more, chromosomes that differ in sequence, are retained less than their corresponding homodupulices and sequence variation is recognized by the appearance of two, or more, peaks in the chromatographs. Numerous SNPs have been identified on the non-recombinant portion of the Y chromosome by using this technique. To investigate the DNA variation within Pakistan 15 Y-SNPs, an Alu insertion, a LINE1 insertion and the 12f2 deletion, mapping on the non-recombining portion of the human Y chromosome, were typed in 834 Pakistanimales. The combination of these biallelic markers identified 11 stable Y chromosomal lineages or Y‘haplogroups’ in the Pakistani population.Haplogroup frequencies were generally similar tothose in neighboring geographical areas and indicate that there was a common pool of Y lineages within Pakistan that are predominantly from West and Central Asia.
https://www.ijbiotech.com/article_6875_5cd0886f59244daa1fd219bfc64e80eb.pdf
DHPLC
Y-Polymorphisms
Y SNPs
Population genetics
Pakistan
eng
National Institute of Genetic Engineering and Biotechnology of Iran
Iranian Journal of Biotechnology
1728-3043
2322-2921
2003-01-01
1
1
31
35
6876
Improved Procedure for Screening Expression Libraries for Novel Autoantigens
MH Sanati
1
C Stanyon
2
F Alasti
3
PR Carnegie
4
National Research Center for Genetic Engineering and Biotechnology (NRCGEB),
Biotechnology Research Group, Murdoch University,
National Research Center for Genetic Engineering and Biotechnology (NRCGEB),
Biotechnology Research Group, Murdoch University,
The standard method for immunoscreening of a cDNA expression library is time-consuming becauseof the production of a large proportion of false positives during the first and second round of screening.This problem is more important when a sensitive chemiluminescence detection system is used. Due tothe high sensitivity of the detection system, there is a need to avoid false positives which occur when theantibody reacts non-specifically. False positives are generally eliminated through absorption of the antibodywith the host bacteria and by eliminating any clones, which react with antibodies present in normalsera. Here we present a method of obtaining almost identical bacteriophage plates by culturing phage inparallel, and show that this technique produces positive plaques in duplicate and eliminates false positives.Using this method, we successfully screened a human fetal spinal cord lambda gt11 cDNA libraryusing purified immunoglobulin G (IgG) from patients with multiple sclerosis (MS) and Guillain – Barre syndrome (GBS).
https://www.ijbiotech.com/article_6876_eeed0b847c39b7a00b9f2f7104ff09a2.pdf
Immunoscreening
Expression
cDNA Library
Auto-antigen
Lambda
eng
National Institute of Genetic Engineering and Biotechnology of Iran
Iranian Journal of Biotechnology
1728-3043
2322-2921
2003-01-01
1
1
36
40
6877
Study of Desulfurization Rate in Rhodococcus FMF Native Bacterium
S Akbarzadeh
1
J Raheb
2
A Aghaei
3
AA Karkhane
4
National Research Center for Genetic Engineering and Biotechnology (NRCGEB)
National Research Center for Genetic Engineering and Biotechnology (NRCGEB),
National Research Center for Genetic Engineering and Biotechnology (NRCGEB),
National Research Center for Genetic Engineering and Biotechnology (NRCGEB),
The Rhodococcus FMF (R. FMF) native bacterium was isolated from soil contaminated with oil in Tabrizrefinery. This bacterium carries three genes sox (dszA, B, C) on its genomic DNA. Preliminary studieshave proved that R. FMF strain possess desulfurization activity. In this work soxA and B genes wereamplified by PCR, after designing a pair of suitable primers. Analysis of PCR products on agarose gelelectrophoresis showed a sharp band in the 2.46 kb region, belonging to soxA and B genes. In addition,dot blotting using the sox4 probe, confirmed the presence of sox operon in the PCR product. The standardGibbs test was designed for desulfurization activity assay in which production of 2HBP (μM) in four bacterialstrains [R. FMF, P. aeruginosa (pESOX4), E.coli DH5α (pESOX3) and E. coli CC118λpir (pESOX4)]was recognized and compared. After 41h of dibenzothiophene (DBT) addition, Pseudomonas aeruginosa(pESOX4) produced 4.8 μM 2HBP and the local R. FMF produced 3.8 μM of DBT. Comparison of 2HBP production in the standard strain of P. aeruginosa (pESOX4) with the isolated R. FMF strain fromTabriz refinery revealed that the later is equally capable of desulfurizing DBT.
https://www.ijbiotech.com/article_6877_c103cca839412e73379807a1a4fe1863.pdf
Desulfurization
Dibenzothiophene
Rhodococcus
eng
National Institute of Genetic Engineering and Biotechnology of Iran
Iranian Journal of Biotechnology
1728-3043
2322-2921
2003-01-01
1
1
41
46
6883
Silver Resistance In Acinetobacter baumannii BL54 Occurs Through Binding to a Ag-Binding Protein
MR Shakibaie
1
BA Dhakephalker
2
BP Kapadnis
3
BA Chopade
4
Department Of Genetics, Research Center, Kerman University of Medical sciences, Kerman, Iran.
Department Of Microbiology, University Of Pune, Pune 114700, India.
Department Of Microbiology, University Of Pune, Pune 114700, India.
Department Of Microbiology, University Of Pune, Pune 114700, India.
The mechanism of plasmid mediated silver (Ag) resistance was investigated in Acinetobacter baumanniiBL54. The intracellular accumulation of Ag in both original strain BL54 and Escherichia coli K12transconjugant containing plasmid pUPI276 began immediately and reached a maximum within 60 minutes.This initial accumulation was followed by net loss of Ag which reached a maximum within 180 min.Pre-treatment of cells with 0.5 mM 2,4 dinitrophenol (DNP); 20 mM N, N-dicyclohexylcarbodiimide(DCCD); 3% toluene; 25 mg/ml cefotaxime and polymyxin-B resulted in considerable decrease in theaccumulation process. Ags plasmid less cured derivative (BL54.1) also accumulated silver but only onefourththe amount compared to the resistant strain BL54. The intracellular accumulated silver is detoxifiedby binding to a cysteine rich metal binding protein.The purified Ag-binding protein exhibited maximum absorption at 280/215 nm. From the above datait could be concluded that the intracellular detoxification of silver in A. baumannii BL54 is achievedthrough binding to a cysteine rich metalloprotein.
https://www.ijbiotech.com/article_6883_b32c011bfde9f2a691da4aa63f8684a8.pdf
Acinetobacter
Silver resistance
Silver accumulation
Metalloprotein
eng
National Institute of Genetic Engineering and Biotechnology of Iran
Iranian Journal of Biotechnology
1728-3043
2322-2921
2003-01-01
1
1
47
58
6878
Genetic and Molecular Dissection of Blast Resistance in Rice Using RFLP, Simple Sequence Repeats and Defense-Related Candidate Gene Markers
Ali Moumeni
alimoumeni@yahoo.com
1
Hei Leung
2
Rice Research Institute of Iran, P. O. Box 1658-41635, Rasht, Iran.
International Rice Research Institute, Manila, The Philippines.
Blast, Pyricularia grisea (Cooke) Sacc., is one of the most destructive diseases of rice worldwide and canresult in significant reductions in yield. The use of resistant cultivars is the most economical and effectiveway of controlling rice blast. A variety of DNA markers, including plant defense-related candidategene markers are available for genetic characterization and molecular analysis of rice. A set of 161recombinant inbred lines, RILs, from a cross between Nemat, an improved and high yielding cultivar, andAnbarboo, a traditional and aromatic rice, was used to identify defense-related candidate gene, RFLPand SSR markers linked to components of resistance to blast, i.e. infection type, lesion density, the percentof diseased leaf area, and lesion size in rice. The RILs were tested using two single blast isolates ingreenhouse, and field population of blast in blast nursery in International Rice Research Institute,Philippines, in 2000–2001. Of the 86 defense-related candidate gene, 153 RFLP, and SSR markers 26defense-related candidate gene, 66 RFLP, and 85 SSR markers were polymorphic in two parental lines.Results showed that a defense gene, b8, a NBS-LRR originated from barley, closely linked to different components of resistance to blast. The defense genes of r5, r7, PrP2, and ERS from rice, maize, andArabidopsis, respectively, have had minor effects on different components of resistance to blast. TheRFLP markers, i.e. RZ536, RG351, RZ76, RZ397 on chromosomes 7, 11 and 12, and the SSR markersincluding RM224, RM179, and RM277 on chromosomes 11 and 12 were tightly linked to componentsof resistance to blast. The linked markers can now be used for resistance gene pyramiding and markerassistedselection in the breeding population. The results suggested the presence of race-specific resistancegenes exhibiting strong differential pathogenhost interaction. We need to incorporate new sources of gene pool to make the genetic base broaden.
https://www.ijbiotech.com/article_6878_c5b91270cab2f35ea62527eceff8382d.pdf
Blast resistance
RILs
RFLP
SSR
Defense related candidate gene
Rice
eng
National Institute of Genetic Engineering and Biotechnology of Iran
Iranian Journal of Biotechnology
1728-3043
2322-2921
2003-01-01
1
1
59
64
6882
PCR optimization: Improving of Human Cytomegalovirus (HCMV) PCR to Achieve a Highly Sensitive Detection Method
S Amini Bavil Olyaee
1
Farzaneh Sabahi
2
Mohsen Karimi
3
Virology Department, School of Medical Sciences, Tarbiat Modarres University, P. O. Box: 14115-111, Tehran, Iran.
Virology Department, School of Medical Sciences, Tarbiat Modarres University, P. O. Box: 14115-111, Tehran, Iran.
Biotechnology Research Center (BRC), Pasteur Institute of Iran, Tehran, Iran.
Polymerase chain reaction (PCR) is a rapid and simple technique with high sensitivity and specificity. Inthe recent years, PCR has been used for rapid detection of viral nucleic acids, such as Humancytomegalovirus (HCMV), whereas, PCR optimization is an important task to be done, especially beforeit’s diagnostic application. Annealing temperature, ion concentration (especially Mg2+ ion) and thecycling program and enhancer compounds are important optimization parameters. Peripheral bloodleukocytes (PBLs) were isolated from samples collected from renal transplant recipients suffering fromsevere and symptomatic CMV disease. PBLs DNA was extracted and used for PCR. Annealing temperatureand MgCl2 concentration and cycling condition were optimized. Dimethyl sulfoxide (DMSO) and gelatinwere checked as enhancer components. The optimized condition obtained through this study was:1x PCR buffer (20 mM Tris-HCl pH 8.6, 50 mM KCl), 2.5 mM MgCl2, 0.2 mM of each dNTPs, 0.25 μM ofeach primers, 0.25 unit/25 μl Taq DNA polymerase, 5% DMSO, 500 μg/ml gelatin and 50-150 ng templateDNA in 25 μl final volume. PCR was performed as: 95°C 5 min (pre-denaturation), 94°C 50 sec, 58°C1 min, 72°C 1 min for 35 cycles and 72°C 5 min (final extension). Using these conditions, it was shown thatoptimized PCR was five fold more sensitive thaninitial PCR; which can be used for diagnostic applicationof HCMV in renal transplant patients.
https://www.ijbiotech.com/article_6882_1ab7870ebe4e87adea66d86f1109596f.pdf
Polymerase chain reaction (PCR)
optimization
Human Cytomegalovirus (HCMV)
Renal transplant recipients