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Volume 4 Special Issue 1 2010
Including
Proceedings of the 2nd International Conference for Applications of Biotechnology
October 17-18, 2009, MSA University, Cairo, Egypt
ISBN 978-4-903313-50-4
How to reference: Feng JX, Stallsmith BW, Davis MR, Podila GK (2010) PCR-enriched Differential Display of Gene Variation. In: Mansour AMMA (Ed). Dynamic Biochemistry, Process Biotechnology and Molecular Biology 4 (Special Issue 1), 1-5
Guest Editor
Ahmed Mansour Mohamed Mansour Alzohairy
Genetics Department, Faculty of Agriculture, Zagazig University, Egypt
http://english.zu.edu.eg/
CONTENTS AND ABSTRACTS
Jason X. Feng, Bruce W. Stallsmith, Maria R. Davis, Gopi K. Podila (USA) PCR-enriched Differential Display of Gene Variation (pp 1-5)
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ABSTRACT
Original Research Paper: Discovery of genetic variation-based biomarkers is time-consuming and cost-prohibitive. We have developed an integrated high-throughput procedure to comprehensively screen and identify differentially expressed genetic variation-based biomarkers and biomarker-related protein-coding genes. These identified biomarkers and marker-genes will be further utilized to analyze their corresponding chromosome variation using reverse genetic method from published data. These biomarkers, the marker-related genes and their genomic variations will play critical roles in clinical diagnosis, drug discovery and pathogenesis research of diseases. In this study, differentially expressed genes between two mRNA samples are discriminately amplified and displayed. Newly discovered special genetic sequence information is analyzed and further used to develop diagnostic biomarkers. This approach can be used to screen biomarkers from cancer genomes, genetic diseases, and for microorganism characterization.
Ali Shakeri-Zadeh, G. Ali Mansoori (USA), A. Reza Hashemian, Hossein Eshghi, Ameneh Sazgarnia, A. Reza Montazerabadi (Iran) Cancerous Cells Targeting and Destruction Using Folate Conjugated Gold Nanoparticles (pp 6-12)
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ABSTRACT
Original Research Paper: We report here the production and use of a new nanoconjugate made up of folate and gold nanoparticle (AuNP) and conjugated by 4-aminothiophenol (4Atp), named Folate-4Atp-AuNP and applied for the selective cancer cells targeting and destruction. Its excellent selective cancer cells targeting capability is due to its folate segment. Its highly localized cell damage capability is because of photothermal treatment due to the presence of the gold nanoparticle, which can strongly absorb visible light and rapidly convert it into heat. The level of light-induced damage and cytotoxicity of the Folate-4Atp-AuNP nanoconjugate is investigated using MTT assay. Experiments are conducted on human adenocarcinoma HeLa cell chosen as the model cancer cell line because of its folate receptor overexpression (Masters 2002). In addition, MCF7 cell line was used as the “blank sample” because of its low level of folate receptor expression (Dickson et al. 1986). Identical Folate-4Atp-AuNP nanoconjugate concentrations, incubation periods and radiation exposure were used for both cell lines. The effect of light–gold nanoparticle interaction on cell lethality is examined and reported in detail. No significant cytotoxicity caused by the Folate-4Atp-AuNP nanoconjugate and cell damage induced by light was observed for both cell lines. Following photothermal treatment, more than 95% cell killing was achieved for HeLa cells whereas cell lethality for MCF7 was up to 26%. It was found that cell lethality depended strongly on incubation period and the nanoconjugate concentration. The cancerous cell targeting and destruction method described in this study is an effective photothermal cancer therapy technique, which can be effective for in vivo cancer therapy.
Kátia Nardelli Greco, Marta Cristina Souza, Laura Araújo Tomé, Rita de Cássia Leone Figueiredo-Ribeiro, Rosemeire Aparecida Bom Pessoni (Brazil) Inulin of Vernonia herbacea from the Brazilian Cerrado Changes Colon Morphology and Lipid Metabolism in Swiss Mice (pp 13-18)
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ABSTRACT
Original Research Paper: The present work describes the influence of inulinobtained from reserve organs of Vernonia herbacea on colon morphology and lipid metabolism in Swiss mice. Animals were fed for five weeks either with Nuvilab CR-1 (control diet) or with Nuvilab CR-1 supplemented with 10% V. herbacea inulin or with 0.5% cholesterol or 0.5% cholesterol + 10% inulin. Distal colon sections were prepared and stained for morphometrical analysis and histochemical characterisation. Enzymatic assays were used to quantify plasma lipids and lipoproteins. The VLDL fraction and triglycerides were estimated by the Friedwald method. The intake of V. herbacea inulin-supplemented diet increased neutral mucins, stimulated sulphomucin production and decreased sialomucin secretion, compared to the control group. It also improved crypt depth and number of goblet cells (P < 0.05 vs. control). HDL-cholesterol concentrations tended to be higher with inulin, as was the ratio of HDL to LDL. V. herbacea inulin has beneficial effects on mouse colon morphology and lipid metabolism.
Irene Axarli, Nikolaos E. Labrou (Greece) Directed Evolution of Cytochrome P450 CYP102A2 from Bacillus subtilis (pp 19-24)
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Original Research Paper: Bacillus subtilis flavocytochrome CYP102A2 is a high activity fatty acid hydroxylase that has evolved from fusion of a eukaryotic-like NADPH-cytochrome P450 reductase (CPR) to a P450 in a single polypeptide chain. In the present work we report the directed evolution of CYP102A2 from B. subtilis with a focus on its substrate specificity. The highly active CYP102A2 was subjected to error-prone PCR (epPCR) to generate enzyme variants with altered substrate specificity. The library of CYP102A2 mutants was expressed in BL21(DE3) Escherichia coli cells and screened for their ability to oxidize several substrates (sodium dodecyl sulphate, lauric acid, 1,4-naphthaquinoline, 2-hydroxy-1,6-naphthoquinone and ε-amino-n-caproic acid) by means of an activity assay. After a single round of epPCR, the variant Pro15Ser/Phe160Leu was isolated which exhibited altered substrate specificity towards naphthoquinones. Molecular modeling of CYP102A2 monooxygenase domain suggests that Phe160 is located at the end of α-helix-6 and is involved in van der Waals interactions with residues positioned at the α-helix-10 which are involved partly in the formation of the substrate binding pocket. Therefore Phe160 seems to affect substrate binding and catalysis indirectly.
Haggag S. Zein (Egypt), Jaime A. Teixeira da Silva, Kazutaka Miyatake (Japan) Sequence Analysis of Monoclonal Antibodies Specific for CMV Coat Protein Subgroup II and Measuring their Affinity Constants with Competitive ELISA (pp 25-31)
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Original Research Paper: Availability of diagnostic methods for plant viruses provides greater flexibility, increased sensitivity, and specificity for rapid diagnosis of virus diseases in disease surveys and epidemiological studies. A total of four hybridomas were generated from BALB/c mice immunized with the m2 strain of Cucumber mosaic virus coat protein subgroup II (CMV-CPII). A modified reverse-transcriptase PCR protocol was used to amplify and sequence the light (VL) and heavy chains (VH) of the V-genes in order to produce monoclonal antibodies (mAbs) with high specific affinity for CMV-CPII. The VH and VL of mAbs produced were characterized and sequenced and compared with the GenBank database. Database analysis of the sequences that encoded the V-genes showed that the light chains of the four hybridomas expressed the family Vκ1A gene, bb1.1, while four of the heavy chains genes expressed four genes of the family VH1/J558, gene VH130. There was frequent addition of random nucleotides of the N region and expected variation in the lengths of CDR3 regions, which form the center of the antigen binding site. Somatic mutation, junctional diversity and alternative light chains collectively impart specificity to these serologically distinct epitopes. Apparent dissociation constants (Kd) of mAbs were determined by indirect competitive ELISA yielding Kd = 2.7-5.2×10-7 M. The serological differentiation of CMV isolates is of importance in breeding for disease resistance and in studying disease epidemiology, so specific mAbs were developed, as one of the goals, to provide tools for serotyping isolates of the virus.
Yaswaree Mihilall, Ackmez Mudhoo, Romeela Mohee (Mauritius) Preliminary Study on Compost Production as Substrate for Pleurotus sajor-caju (Fr.) Singer Mushroom Cultivation in Mauritius (pp 32-43)
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Original Research Paper: The in-vessel composting of a mixture of three potential substrates, namely banana leaves (B), mixed cardboard wastes (C) and green Stenotaphrum dimidiatum grass (G) was studied for three weeks. The first compost mix ratio monitored was 16 kg banana leaves, 8 kg cardboard wastes and 16 kg Stenotaphrum dimidiatum (wet basis). The initial wet moisture content, initial porosity and initial wet bulk density for the BCG mix were 69.5%, 86.9% and 152.7 kg/m3, respectively. The peak temperature recorded for the BCG mix at day 10 was 55.1°C. The temperature then decreased to 35 to 45°C for a week. It was noted that there was an aggressive colonisation of an unidentified mushroom mycelium on the BCG mix. Mycelium and pinheads thrived mostly on the surfaces of the cardboard mix and throughout the feedstock, where temperatures averaged 35°C and less. The feedstock was removed from the reactor and mixed with 14 kg banana leaves, 14 kg grass (Stenotaphrum dimidiatum) and 1.5 L water (second BCG mix), and the composting experiment run afresh. A peak temperature with a maximum of 60.7°C was noted on day 6 and no mycelium growth was observed. It was hence inferred that temperatures should reach a minimum of 60°C to produce a sterile compost substrate for the growth of the oyster mushroom mycelium. The final parameters of the compost after 17 days were as follows: wet bulk density 271 kg/m3, wet moisture content 54.3%, total dry solids 34.7%, net estimated volatile solids (VS) decrease of 40.5% (fixed ash basis), pH 7.5, electrical conductivity 1.57 mS/cm and respiration rate 2.52 mgCO2.C/day.gVS.
Supriyo Basak, Meenakshi Kumari, Amrita Kumari, Prasanna Jyothi D (India) Production of Cellulase from Myrothecium verrucaria NCIM 903 by Solid State Fermentation Utilizing Wheat Bran as Substrate (pp 44-45)
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Research Note: In the present work, an attempt has been made to produce fungal cellulase enzyme from Myrothecium verrucaria NCIM 903 by solid substrate fermentation. Potato dextrose agar was taken as a seed medium for maintenance of the culture. Substrate used for solid substrate fermentation was wheat bran. Total cellulase activity was analyzed by filter paper assay and the maximum cellulase activity of 0.23 U/g was found after 3 days of fermentation.
Tarek M. Salman, Mohamed M. El Zahaby, Ossama A. Mansour, Gamal A. Omran, Soad M. Gomaa, Hisham S. Gad (Egypt) Effect of Narcotic Addiction on Hypothalamic Pituitary Gonadal Axis Hormones (pp 46-49)
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Original Research Paper: Drug addiction is considered a serious societal problem, often leading to social maladaptation, decreased work productivity and job loss. Beside loss of control, drug addiction gradually leads to the development of tolerance and a demand to take higher doses in order to attain the desired stimulating effect. This study intended to investigate the effect of bhang (which consists of the leaves and flowering tops of male and female Cannabis sativa plant) and opium addiction on hypothalamic pituitary gonadal axis hormones. It was conducted on 83 individuals whose ages ranged from 23 to 35 years and who were classified into 6 groups. Group A: male subjects addicted to opium; group B female subjects addicted to opium; group C male subjects addicted to bhang; group D female subjects addicted to bhang; group E control male subjects and group F control female subjects. Blood sampling from female groups (addicts and control) were taken during the follicular phase. The results of our research revealed a significant decrease in serum total testosterone, follicle stimulating hormone (FSH), leutinizing hormone (LH) and prolactin in male addicts to opium in relation to the control group. Although the serum level of these hormones was significantly lower in males addicted to bhang when compared to the control group, the effect of opium was relatively more severe than the effect of bhang. However, the decrease in serum estradiol in male addicts was not significant when compared to the control group. Moreover, the results showed a significant decrease in serum testosterone, estradiol, FSH, LH and prolactin levels in female addicts to opium more than the female addicts to bhang when compared to the control group except for estradiol, for which the effect of bhang was more severe.
Tarek M. Salman, Mohamed M. El Zahaby, Ossama A. Mansour, Gamal A. Omran, Soad M. Gomaa, Hisham S. Gad (Egypt) Oxidative Stress and Lipotoxicity of Bhang and Opium Addiction. Effects on Adrenal Gland Secretions (pp 50-54)
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Original Research Paper: This study was carried out to investigate the effect of bhang and opium addiction on hypothalamic pituitary adrenal axis hormones and the role of free radicals in lipotoxicity induced by bhang and opium addiction. 83 individuals, whose ages ranged from 23 to 35, were classified into 6 groups: Group A, male subjects addicted to opium; group B, female subjects addicted to opium; group C, male subjects addicted to bhang; group D, female subjects addicted to bhang; group E, control male subjects; group F, control female subjects. Blood sampling from female groups (addicts and control) were taken during the follicular phase. Plasma and serum were separated to determine total cholesterol, triacylglycerol (TAG), HDL- and LDL-cholesterol, serotonin, dopamine, adrenaline, catalase, thiobarbituric acid-reactive substances (TBARS), protein oxidation and protein thiols. There was a significant increase in serum total cholesterol, TAG and LDL-cholesterol in male opium and bhang and female opium and bhang addicts, respectively in relation to the control group. However, the decrease in serum HDL-cholesterol in male and female addicts was not significant when compared to the control group. Moreover, there was a significant increase in serotonin (258.4 ± 0.692, 260.2 ± 0.734 and 259.5 ± 0.698, 259.8 ± 0.705), dopamine (86.95 ± 1.935, 82.66 ± 2.287 and 88.01 ± 2.108, 82.94 ± 2.360) and adrenaline (55.87 ± 2.212, 56.93 ± 1.830 and 57.57 ± 1.500, 58.40 ± 1.635) levels in male opium and bhang and female opium and bhang addicts, respectively compared to the control male (213.3 ± 1.483, 54.37 ± 1.432 and 36.67 ± 1.025) and control female (212.9 ± 1.445, 55.42 ± 1.320 and 35.83 ± 1.029) group, respectively. In addition, there was a significant increase in oxidative stress markers, TBARS, and protein oxidation and a decrease in antioxidants, catalase and protein thiols in male opium and bhang and female opium and bhang addicts, respectively in relation to the control group.
Gamal Saadi, Mervat El-Ansary, Samah Abd El-Hamid, Iman Abd El-Aziz (Egypt) Mesenchymal Stem Cell Transfusion as a Novel Immunosuppressive Regimen with Possible Induction of Microchimerism (pp 55-60)
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Original Research Paper: Human mesenchymal stem cells (MSCs) have immunosuppressive capacities. Although their efficacy is currently studied in graft versus host disease (GVHD), their effect on alloreactivity in solid organ transplant (SOT) patients is unknown. Our work aimed to use allogeneic donor-specific MSCs (DS-MSCs) transfusion prior to renal transplantation as an immunosuppressive induction regimen. Our study included 4 groups of patients, all of which were diagnosed with chronic renal failure and had undergone renal transplantation. The first group included 7 patients that were induced by DS-MSCs. The second included 6 patients induced by antithymocyte globulin (ATG). The third included 6 patients induced by anti-CD25 while the 4th group included 7 patients who received no induction. The immunosuppressive regimen was cyclosporine (CsA), Mycophenolate mofetil (MMF) and prednisolone (PRD) for all patients. Bone marrow (BM) (90 ml) were aspirated from the iliac bone of related donors, to separate MSCs, then about 10 million MSCs placed in 10 ml saline were infused intravenously in 2 divided doses 1 week apart. Our results showed that the lowest mean serum creatinine level measured after 1, 3, and 6 months were in those patients who received pre-transplantation DS-MSC infusion (group I). Also rejection was less frequent in patients of group I. Microchimerism was detected after MSCs transfusion in one case of group I. We conclude that MSCs can escape immune recognition, can inhibit immune responses and prevent the development of cytotoxic T-cells so their transfusion may be used to treat organ allograft rejection and reduce the need for an immunosuppressive regimen after renal transplantation.
Mervat El-Ansary, Mervat Mattar, Shereen Kamel, Samah Abd El-Hamid, Rania Abou Elnour (Egypt) Dendritic Cell Vaccine as an Adjuvant Therapy in the Treatment of Chronic Myeloid Leukemia (pp 61-66)
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Original Research Paper: Dendritic cells (DCs) are bone marrow (BM)-derived antigen-presenting cells (APCs) with a key role in the induction of immunity. This study aimed at generating a DC vaccine expressing leukemia-associated antigen differentiated from myeloid blast cells to boost the immune system and to improve the clinical outcome of chronic myeloid leukemia (CML) patients. Two ml of venous blood were obtained from each patient to generate DCs by suspending them in liquid culture medium containing granulocyte monocyte colony stimulating factor (GM-CSF) and interleukin 4 (IL-4) and activated by adding tumor necrosis factor alpha (TNFα). DCs, identified by CD83 expression using flow cytometry, showed a significant increase after culture. A follow-up of patients by monitoring the immunological response was done by flow cytometric assessment of CD8+ T-cells % before and after injection of DCs. This study included 22 patients diagnosed as chronic-chronic myeloid leukemia who were divided into 2 groups. Group I was a pathological control group which included 13 age- and sex-matched CML patients that were not given the DC vaccine and were injected with saline. Group II included 9 CML patients that were given the DC vaccine. The WBC count before (141.59 ± 100.51) and after (94.47 ± 76.14) vaccination was highly significantly different in group II patients (P = 0.018). Although there was no statistically significant difference in the median of the CD8+ level and the absolute number of CD8+ before and after vaccination, there was an actual increase in the percentages of both medians. We conclude that a DC vaccine may be used as an adjuvant therapy parallel to CML patients’ chemotherapeutic regimen.
Waleed Nazmy El Mazney, Mohamed Reda Mohamed Diab, Yasmine Mamdouh William (Egypt) Multifunctional Implantable Chitosan for Skin Tissue Engineering and Wound Healing (pp 67-73)
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Original Research Paper: Chitosan, the deacetylated derivative of chitin, is considered as a new technology for treatment of skin wounds. In this study, primary cell cultures of baby mice (1-2 days) skin tissue, supplemented with 10% fetal calf serum were seeded onto air-dried chitosan. In a mice model, the cells developed to epithelial tissues can be used for implantation for skin regeneration and wound healing. Under anesthesia, wounds of the same size were induced in each tested mouse and their chitosan-free control counterpart. Tested mice included two chitosan-treated groups, one with a suture wound while the other was left with an open wound. Assessment of the treatment efficacy revealed the following. On day 9 the wound closure of the suture chitosan group was better than the suture control group. Despite this, the closure rate of the suture control group continued to increase and became nearly equal at day 11 to that of the suture chitosan group, showing a significant increase on days 13 and 16 compared to the test group. For the open wound groups, on days 3 and 9 the wound closure of both the chitosan and control groups were similar, but on days 11 and 13 the chitosan group significantly accelerated wound closure and completely healed on day 16 compared to the control and sewn groups. Results clarify the ability of using a biodegradable implement in transplanting culture substrates for skin regeneration in an open wound.
Thanaa E. Helal, Monir A. El-Ganzuri, Ahmed M. Aref, Mohamed T. Badr (Egypt) Expression of p53 Protein in Hepatocellular Carcinoma: Relationships to Viral Infection and Prognostic Factors (pp 74-78)
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Original Research Paper: Hepatocellular carcinoma (HCC) is very common in Egypt due to the high incidence of hepatitis C virus (HCV) infection. The aim of the current study was to assess the relationships between p53 and the viral infection as well as the prognostic factors in patients with HCC. p53 protein was examined by means of immunohistochemistry in tissue sections from 34 HCCs and 16 cirrhotic patients. p53 was expressed in 12 out of 34 HCCs (35%). All the 16 cirrhotic cases were p53-negative. No significant relationship was detected between p53 expression and the prognostic factors(P > 0.05). However, the level of p53 expression was significantly higher in patients coinfected with HCV and hepatitis B virus (HBV) than in those having single infection.
Ahmed Mansour (Egypt), Jaime A. Teixeira da Silva (Japan), Eman E. El-Araby (Egypt) Molecular Markers Associated with the Development of New Phenotypes of Japanese Quail in Egypt (pp 79-84)
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Original Research Paper: In this report, genomic variation within four isolated phenotypes of Japanese quail (Coturnix japonica) in Egypt were investigated using two different molecular marker systems which target different genomic regions: RAPD (random amplified polymorphic DNA) and ISSR (inter-simple sequence repeat). Different dendrograms were constructed based on RAPD and ISSR results, individually and collectively, revealing that similarity and clustering is highly dependant on the marker system used.
Hassan Moawad, Osama M. Darwesh, Wafaa M. Abd El-Rahim, Olfat S. Barakat, Mohamed Z. Sedik (Egypt) Evidence of Biodegradation of Reactive Red Textile Azo Dye in an Anoxic/Aerobic Bioremediation System (pp 85-90)
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Original Research Paper: The biodegradation of reactive red (RR) textile azo dye was investigated. Three bacterial strains, Enterobacter cloacae, Pseudomonas sp. and Bacillus sp. were used to decolourize and/or degrade the dye. A small bench-scale suspended bed bioreactor was used to study the capacity of these three bacterial strains to decolourize the dye solutions added as 5 successive supplements. The degradation of azo dyes was judged by the formation of aromatic amines (first product of dye biodegradation). All bacterial strains tested in this study produced aromatic amines under anoxic conditions. Additional evidence for biodegradation of RR textile azo dye was determined using 3 bioassay methods. Six test strains contributing to soil fertility were grown in spent media obtained from RR biodegradation. The growth of each test strain on biodegradation products was as high as the growth on the same specific media for each strain. This is evidence for the removal of toxicity in the biodegradation products. Another bioassay method used the germination of plant seeds to test dye phytotoxicity. The biodegradation of RR removed the phytotoxic effects of the dye on wheat (Triticum aestivum Giza ‘167’) and berseem clover (Trifolium alexandrinum L.) seed germination. The chemical oxygen demand (COD), which indicates the organic load in the medium, decreased from 2048 to 599 ppm under aerobic conditions. The bacterial growth after biodegradation of RR increased indicating the breakdown of organics. A continued decrease in the COD value indicates a steady biodegradation of the anoxic biodegradation products as the growth conditions turned aerobic. This study shows that the biodegradation of a toxic textile azo dye, RR, is possible by using potent bacterial strains in a sequential anoxic/aerobic bioremediation system.
Elsayed A. Fayzalla, Mohamed E. Abdalla, Mona G. Zaghloul, Sahar R. Gedara, Elsherbiny A. Elsherbiny (Egypt) Antifungal Potential of Extracellular Metabolites Produced by Drechslera spp.against Phytopathogenic Fungi (pp 91-99)
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Original Research Paper: Culture filtrates of nine Drechslera isolates (D. australiensis, D. cactivora, D. cynodontis, D. ellisii, D. hawaiensis, D. maydis, D. neergaardii, D. poae and D. spicifera), used at 30, 50 and 70%, were evaluated in-vitro against mycelial growth and spore germination of eight plant pathogenic fungi (Alternaria solani, Botrytis cinerea, Botrytis fabae, Fusarium oxysporum, Fusarium solani, Rhizoctonia solani, Sclerotinia sclerotiorum and Sclerotium cepivorum). Among the tested culture filtrates, only D. cynodontis was a highly effective growth inhibitor against all tested fungi; it reducing the fungal growth from 51.1 to 86.7%, and was the strongest inhibitor of spore germination, inhibiting spore germination of all tested fungi by 92 to 98%. The chloroform extract of D. cynodontis was the best growth inhibitor against all tested fungi: it inhibited fungal growth from 66.7% to 88.9% at 30 mg/ml and also highly effectively suppressed spore germination of all tested fungi at all concentrations. In greenhouse experiments, the chloroform extract was most effectively controlled damping-off disease caused by F. solani and S. sclerotiorum on bean. The ethyl acetate extract was the second best for S. sclerotiorum and R. solani. Two sesquiterpenes, dihydrobipolaroxin (1) and sorokinianin (2), were isolated from the chloroform extract of D. cynodontis and identified by EI-MS, 1H-NMR, 13C-NMR, DEPT, COSY, HMQC and HMBC experiments. Compound (1) was a highly effective growth inhibitor against A. solani, F. oxysporum and S. sclerotiorum at 100 µg/ml. Compound (2) decreased the growth of all tested fungi between 22.2 and 61.1%. Compound (1) greatly decreased spore germination of A. solani and F. solani. Compound (2) highly effectively suppressed spore germination of F. solani at 100 µg/ml.
Khaled Adly Mohamed Khaled (Egypt), Jaime A. Teixeira da Silva (Japan) Molecular Profiling Using Protein Markers for Salt Tolerance in Sugarcane (pp 100-103)
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Original Research Paper: This study was carried out between 2006 and 2008 with the aim of evaluating five sugarcane varieties (G.84-47, G.98-28, G.95-21, G.95-19 and Phil8013) for their tolerance to salinity stress in comparison with the commercial cultivar G.T.54-9; SDS-protein analyses were conducted to identify protein markers associated with salinity tolerance for selection of promising lines tolerant to salt stress at an early stage of the sugarcane breeding program. The performance of varieties was assessed in sand culture using salinized (8000 ppm NaCl) nutrient solution under greenhouse conditions. G.95-21 and G95-19 had superior salt tolerance to salt for most yield-related traits, while commercial varieties G.T.54-9 and Phil8013 were more sensitive. Two SDS-protein markers (MW = 32 and 85 kDa) were positively associated with salt tolerance, since they were identified as being expressed exclusively in the resistant varieties (G.95-21 and G95-19) as a result of the stress.
Khaled Adly Mohamed Khaled, Mohamed Abd-El-Kareem Fathalla (Egypt) Molecular Profiling and Genetic Diversity in Sugarcane using RAPD (pp 104-106)
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Research Note: The genetic diversity of six sugarcane Saccharum sp. genotypes (G.T54-9, G.84-47, G.98-28, G.95-21, G.95-19 and Phil8013) was assessed by detecting polymorphisms with 24 RAPD primers. The genetic similarity between genotypes was assessed on the basis of the Dice similarity coefficient and complemented with a UPGMA-based cluster analysis. Only three primers produced polymorphic amplification products: OP-A01, OP-A07 and OP-B07. A total number of 130 amplified fragments were obtained with the three polymorphic RAPD primers for an average of 43 bands per primer. These polymorphic bands can be used as positive or negative molecular markers for °Brix, sucrose, sugar recovery, cane yield and sugar yield. The six sugarcane genotypes were divided into four clusters. Genetic diversity was lowest between G.T. 95-21 and G 95-19 (two sister lines) but highest between G.T.54-9 and Phi1 80-13 (two commercially grown genotypes). The RAPD-derived genetic similarity indices ranged from 10% between G.T.54-9 and Phi1 8013 to 87% between G.T. 95-21 and G 95-19. By capturing the closeness of the two sister lines and grouping Phil 8013 (derived from a different breeding program) as an outcast, these three primers can provide an additional discriminatory power for genetic diversity and crossing of the working germplasm in our breeding program.
Ibrahim M. Khater (Egypt) Homology Modeling of NS3 Protease of Hepatitis C Virus Genotype 4a (pp 107-108)
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Research Note: The amino acid sequence of the NS3 protease of hepatitis C virus genotype 4a was obtained from the SWISS-PROT database. The tertiary structure of NS3 protease of hepatitis C virus genotype 4a was modeled using Swiss-model server (i.e. Automatic homology modeling server). The percentage identity was > 86%. The model was validated using the Procheck program. The results indicated that at least 89% of the amino acid residues are in the core region and 0% in the disallowed region. |
