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Research Article
Characterization and spatial expression pattern of a novel CONSTANS gene (GbCOL-6) from Ginkgo biloba
Mingyue Fu, Jiaping Yan, Lanlan Wang, Xian Zhou, Junping Tan and Feng Xu
Background and Objective: Ginkgo biloba is an important medicinal plant but the transitional period from the sowing to the flowering of G. biloba is too long, which severely limits its reproduction and application. The CONSTANS (CO), is one of the important transcription factors in the transition from vegetative growth to reproductive growth. The CO plays a central role in photoperiodic flowering control. The aim of this study was to clone and characterize a CONSTANS-like 6 (GbCOL-6) gene from G. biloba. Materials and Methods: A pair of specific primers were designed based on the data of the G. biloba transcriptome. The full-length cDNA of the CO gene was amplified from the total RNA of G. biloba leaves by RT-PCR, named GbCOL-6 (GenBank Accession No. MG251395). The expression of GbCOL-6 gene in different tissues was studied by quantitative RT-PCR. Results: Sequence analysis results showed that the cDNA of GbCOL-6 contained a 1479 bp open reading frame (ORF), which encodes a 492 amino-acid protein. The theoretical molecular weight and pI of the GbCOL-6 are 54.65 kDa and 5.08, respectively. Similar to other CO proteins, GbCOL-6 contains two conserved domains (B-box and CCT domain) and the amino acid sequence showed high similarity to other plant CO proteins. Phylogenetic tree analysis revealed that GbCOL-6 belonged to the second group members of the CONSTANS gene family, which has aB-box domain and a CCT conserved domain. These results indicated that GbCOL-6 is a member of the CONSTANS family and belongs to CONSTANS-LIKE 6. Tissue expression profile analysis showed that GbCOL-6 was expressed in all tissues, the highest expression level was detected in the female strobili and the lowest in the roots. Conclusion: This study found that GbCOL-6 isolated from G. biloba was specifically expressed in the female strobili of G. biloba. However, GbCOL-6 rarely was expressed in roots of G. biloba. This laid the foundation for further understanding of the molecular regulation mechanism of the flowering of G. biloba and shortening the juvenile phase of woody plant through the transgenic technology and providing genetic resources for promoting early flowering.
Research Article
Potential Effects of Taif’s Punica granatum L. Extract on Peripheral Blood Mononuclear Cells from Patients with Rheumatoid Arthritis via Regulation of the Nf-κB Signaling Pathway by the IκBα Gene
Sarah Albogami and Ayman Alhazmi
Background and Objective: Rheumatoid arthritis (RA) is a chronic, inflammatory autoimmune disease characterized by joint inflammation and cartilage and bone destruction. Abnormal nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) signaling contributes to the pathogenesis of RA. The IκBα gene plays an important role as an inhibitor of NF-κB and proinflammatory gene expression. We investigated the expression of the IκBα gene in peripheral blood mononuclear cells (PBMCs) from patients with RA, after treatment with Taif P. granatum L. (TPG) extract. Materials and Methods: The PBMCs from RA patients were treated with 50,100 and 150 μg mL–1 TPG and their RNA was extracted after 1, 2, 4, 6, 8 or 18 h. the IκBα mRNA expression in the samples, determined by polymerase chain reaction, was compared with that in controls, using nonparametric one-way analysis of variance. Results: Dimethyl sulfoxide treatment induced no significant change in IκBα gene expression as compared with that in non-treated control cells; however, there was a significant, dose-dependent increase in IκBα gene expression in PBMCs treated with 50, 100 or 150 μg mL–1 TPG as compared with that in control cells (p<0.0001-0.05). Evidently, the potential effect of TPG extract on PBMCs from RA patients can be detected by changes in IκBα gene expression. Conclusion: The TPG extract markedly increases IκBα gene expression, which might subsequently inhibit NF-κB activation. Thus, TPG extract could be developed as new, safer drug with fewer side effects for RA treatment.
Research Article
Sugarcane Genotypes Assessment Under Drought Condition Using Amplified Fragment Length Polymorphism
Khaled Adly Khaled, Nagwa Ibrahim El-Arabi, Nevien Mahmoud Sabry and Sheren El-Sherbiny
Background and Objective: Improving tolerance of crop plants to different types of environmental stress is an important key to crop production sustainability. Sugarcane productivity and geographical distribution are affected with drought stress. This study was focused on evaluation the performance of eight sugarcane (Saccharum officinarum L.) genotypes under drought stress. Materials and Methods: Eight sugarcane genotypes were assessed for drought tolerant using following yield-related traits:stalk height, stalk diameter, stalk weight, leaf area and number of stalks/plant. Eight AFLP combination were used to detect the genotype specific marker. Results: The eight sugarcane genotypes were assessed for their water stress tolerance in sand culture experiment. Analysis of variance showed significant differences for these traits among the eight genotypes under control and drought treatments. The results indicated that genotypes Co. 285, Co. 997 and Bo.19 were the most tolerant and genotypes Co. 775, F.141 and Co. 396 were the most sensitive ones. However, genotypes G2003-47 and G2007-61 were moderate drought tolerant genotypes. Among the sensitive genotypes, the Co. 775 was the most sensitive one that recorded the highest reduction (%) with all traits except stalk diameter which increased for all genotypes. The Co. 775 and Co. 997 genotypes were used in AFLP analysis. Eight AFLP primer combinations were used to estimate the level of polymorphism among drought tolerant sugarcane genotype Co. 997 and drought susceptible genotype Co. 775. The eight AFLP primer combinations amplified a total number of 886 amplicons, where 55 were polymorphic representing 6.2% polymorphism. Conclusion: The eight genotypes genetically different in their response to drought tolerance. The AFLP marker can be used as genetic marker to assess the sugarcane genotypes and Co. 775 was more sensitive and Co. 997 was most tolerant genotype.
Research Article
Bacterial Diversity Change in Oil-contaminated Soils in Jianghan Oilfield via a High-throughput Sequencing Technique
Lu Gan, Jiang-Ping Wang and Qiang-Ping Wu
Background and Objectives: Oil pollution has become a severe problem in soil ecological in Jianghan Oilfield. The soil bacteria community plays a huge role in maintaining soil ecosystem balance and soil biological activity. The main objective of present study was to examine the change bacterial diversity in oil contaminated soil using a high-throughput sequencing technique. Materials and Methods: The Illumina MiSeq high-throughput sequencing technique was used to study the abundance, diversity and structural change in the oil-contaminated soil in Jianghan Oilfield. Results: The results showed that the abundance and diversity of the soil bacterial community significantly declined under oil pollution. Some dominant species (i.e., Proteobacteria and Actinobacteria) existed in the petroleum-contaminated soils and most were oil-associated and hydrocarbon degrading bacteria. Conclusion: On the basis of this study, the construction of the microbial flora of petroleum degradation is the focus of future research which means a more efficient degradation of petroleum in Jianghan Oilfield.
Research Article
18s rDNA Sequence Analysis of Microfungi from Biofloc-based System in Pacific Whiteleg Shrimp, Litopenaeus vannamei Culture
Nor Azman Kasan, Nurarina Ayuni Ghazali, Nurul Fakriah Che Hashim, Iswadi Jauhari, Ahmad Jusoh and Mhd Ikhwanuddin
Background and Objectives: One of the major problem in Pacific Whiteleg shrimp, Litopenaeus vannamei farming industry is the accumulation of excess toxicant such as ammonia and nitrite. Biofloc technology served as an alternative solution to solve this problem by recycling the water for sustainable aquaculture management due to water scarcity, bio-security and bio-economy. Biological communities within biofloc are mainly composed of micro-organisms which plays an important role in biodegradation of organic material and removal of toxic contaminants. For the purpose of this study, molecular identification of microfungi isolated from biofloc was conducted. Materials and Methods: Isolation of microfungi composition was conducted by culturing the biofloc samples onto Potato Dextrose Agar (PDA) media which further identified molecularly using 18s rDNA sequences analysis. Results: The identified microfungi includes genus from Aspergillus versicolor, A. niger, A. tamarii, A. flavipes, A. aculeatus, Penicillium citrinum, P. griseofulvum, Trichoderma virens and Pestalotiopsis microspora. The most dominant microfungi were genus Aspergillus sp. followed by the presence of Penicillium sp. Trichoderma sp. and Pestalotiopsis sp. Conclusion: Existence of microfungi species such as Aspergillus sp. and Penicillium sp. in biofloc were perfect candidate in forming biofloc that could reduce water pollution towards a sustainable aquaculture industry.
Research Article
Molecular Characterization of Two New Mycoplasma Species Isolated from Chickens in Saudi Arabia
Magdy Hassan Yassin, Alaa Ahmed Mohamed, Mohamed Mahmoud Hassan, Ahmed Abd El-Aziz Baiomy and Amena Maher Ibrahim
Background and Objective: Avian Mycoplasma infections pose a problem in poultry breeding. The main objective in this study was to isolate and identify new endemic Mycoplasma species that cause poultry diseases in Taif, Saudi Arabia by using both traditional microbiological methods and modern molecular methods. Materials and Methods: One hundred forty lung tissue samples from native chickens were collected from a Taif poultry abattoir and subjected to culture isolation, pathological and molecular characterization. Results: Sixteen isolates (11.4%) were identified as Mycoplasma species based on their cellular, morphological and biochemical characteristics. Furthermore, Mycoplasma isolates were identified by 16S rRNA gene sequencing; seven Mycoplasma isolates were identified as Mycoplasma gallinaceum, which infects turkeys. Only one of these isolates was identified as Mycoplasma fermentans, which infects human and eight isolates were unknown Mycoplasma sp. They may be trans located between farmers and chickens or between turkeys and chickens in the same farm. This is the first report of isolation M. gallinaceum from native chickens in the Taif region, Saudi Arabia. Conclusion: These results confirmed that polymerase chain reaction could be used to rapidly diagnose and identify Mycoplasma and is sensitive to molecular detection. These obtained isolates could be applied in control strategies and vaccine development in Saudi Arabia.
Short Communication
Optimization of Allele Specific PCR for the Development of Human Mitochondrial DNA Typing Method
Seri Mirianti Ishar and Rus Dina Rus Din
Background and Objective: Human DNA is categorized into nuclear DNA (nDNA) and mitochondrial DNA (mtDNA). Mutation occurred in these two types of human DNA can be utilized for the purpose of medical reports, molecular genetic analysis, haplogroup variations, forensic crime search and many others. The polymorphisms were selected from mtDNA in both coding and control regions. Main objective of this project was to produce a simple, cost effective yet robust typing method for human identification using allele specific PCR (asPCR) specifically designed for southeast Asian population. Materials and Methods: A total of 60 subjects with ethical approval were randomly collected with 30, 20 and 10 of them represented Malay, Chinese and Indian population. Polymerase Chain Reaction (PCR) was carried out using 25 sets of primers, that amplify the fragments bearing the selected SNPs. In the second round PCR, two types of allele specific primers labelled as wild type allele specific primer (wtASP) and variant type ASP (vtASP) were used to amplify both Cambridge Reference Sequence (CRS) and variant sequence. The PCR was used for first round and for the second round, the as PCR was applied. Finally, the amplified products were directly viewed using UV light. Results: Overall, out of 30 selected SNPs, the designed ASP managed to amplify only 20 SNPs. The selected SNPs in this study were SNP 146, 195, 1709,1719, 1872, 3705, 3027, 3552, 4491, 7684, 9080, 8440, 13626, 16108, 16291, 16274, 16355, 16093, 16335 and 16148 that reported to belong to macrohaplo group M, B, F, E and N. All the amplified products of selected mtSNPs were observed in wild type lane except for SNP 195 that the amplified product was observed in variant lane. Conclusion: The variant allele managed to be amplified with simple technique yet robust to be brought on site. Generally, mutation found using this technique may narrow down the individual and also population hence it is beneficial in cases such as forensic crime and mass disaster.
Research Article
Corrigendum: Size Enlargement and Shorten Longevity of Hibiscus Flower Affected by Gibberellic Acid and Aluminium Sulphate Using Dripping Technique "Published in Biotechnology 13(2): 61-67, 2014"
Mohammed Saifuddin, Mohammad Moneruzzaman Khandaker, Md. Sarwar Jahan, Nashriyah Binti Mat and A.B.M. Sharif Hossain
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