Identification and Characterization of Genes Responsible for Drought Tolerance in Rice Mediated by Pseudomonas fluorescens

Drought is one of the major abiotic stresses which adversely affect crop plants limiting growth and yield potential.Structural and functional characterization of drought stress-induced genes has contributed to a better understanding of how plants respond and adapt to the drought stress.In the present study,differential display technique was employed to study the gene expression of rice plants at the reproductive stage that were subjected to drought stress by withholding water,Pseudomonas fluorescens strain(Pf1) treated plants subjected for drought stress by withholding water and control(well-watered).Differentially expressed c DNAs of six genes(COX1,PKDP,b ZIP1,AP2-EREBP,Hsp20 and COC1) were identified,cloned and sequenced.Real-time q PCR analysis showed that all the six genes were upregulated in drought-stressed plants treated with Pf1.This revealed that the remarkable influence of Pf1 colonization leads to drought tolerance at the reproductive stage.These results showed that high levels of gene expression in plants lacking adequate water can be remarkably influenced by Pf1 colonization,which might be a key element for induced systemic tolerance by microbes.

Quantitative detection of Cymbidium mosaic virus by real time PCR

The technique of SYBR Green-based quantitative real-time reverse transcription polymerase chain reaction(real-time RT-PCR)was applied to quantitative detect a 764 bp nucleotide sequence containing total coat protein(cp)gene of Cymbidium mosaic virus(CyMV).The plasmid containing the target sequence was constructed to prepare the standard curve and detect the sensitivity.The standard curve was drawn based on the linear relationship between the logarithm(base 10)of the quantity of target sequence and cycle threshold[C(T)].While the concentration of plasmid DNA falling within the range of 2.6×10^(7)to 2.6×10^(2)copies per tube established a regression equation,y=-0.3583x+10.32,and related coefficient:r^(2)=0.995.The real-time RT-PCR assay for CyMV had a minimum detectable quantity of two copies per tube.The naturally infected samples of Phalaenopsis sp.and the artificially inoculated samples of Arachnis sp.with trace CyMV were quantitatively detected using this method.CyMVin the positive samples of Phalaenopsis sp.and Arachnis sp.was confirmed by DNA sequencing and cp gene homeology blast.The results showed that CyMV extracted from the leaves of orchid in Hangzhou,Zhejiang Province,China,could be derived from Kunming city(KM),Yunnan Province,China.This method characterized by high sensitivity,specificity,and precision is suitable for early diagnosis and quantitative detection of CyMV.

Human dental pulp stem cells express many pluripotency regulators and differentiate into neuronal cells

Stem cells were isolated from human dental pulp using an optimized method, in which pulp pieces were digested by enzymes and immobilized to enhance cell outgrowth. Stem cell marker expression was detected by reverse transcription-PCR （RT-PCR）, and differentiation markers were detected by real-time quantitative RT-PCR and immunocytochemistry. Results showed that dental pulp stem cells actively expressed nanog, oct4, nucleostemin slain-l, jmjdla, jmjd2c, and cyclin DI. When stem cells were induced to differentiate into neurons, nucleostemin, nanog, and cyclin D1 expression significantly decreased, whereas expression of neuronal markers, such as microtubule associated protein-2 and neurofilament-heavy, significantly increased. These results suggested that stem cells exited a pluripotent state and entered a neuronal differentiation pathway. In addition, results demonstrated that human dental pulp serves as a reservoir of stem cells that express defined stem cell markers; these cells were easily isolated and were induced to differentiate towards a desired cell lineage.