RT-PCR

Overexpression of type-A rice response regulators, OsRR3 and OsRR5, results in lower sensitivity to cytokinins

X. Cheng, Jiang, H., Zhang, J., Qian, Y., Zhu, S., and Cheng, B., Overexpression of type-A rice response regulators, OsRR3 and OsRR5, results in lower sensitivity to cytokinins, vol. 9, pp. 348-359, 2010.

Response regulators are part of a two-component regulatory system. The type-A Arabidopsis response regulators act as negative regulators. To understand the function of type-A response regulators in the model monocot plant, rice (Japonica cultivar-group: Zhonghua11), we overexpressed two type-A OsRR genes, OsRR3 and OsRR5 (pACT1:OsRR3 and pACT1:OsRR5). We hoped to gain insight into their molecular function in cytokinin-signaling pathways.

Differential diagnosis of active hypodermal and hematopoietic necrosis virus based on gene choice and reverse transcription coupled with PCR

M. A. Teixeira, Cruz, J. E. F., Vieira, P. R. N., Branco, I. R. C., Costa, F. H. F., and Rádis-Baptista, G., Differential diagnosis of active hypodermal and hematopoietic necrosis virus based on gene choice and reverse transcription coupled with PCR, vol. 9, pp. 2025-2031, 2010.

The Pacific whiteleg shrimp Litopenaeus vannamei (Penaeidae) is one of the most important cultivated species in world aquaculture. In Brazil, the northeastern states are home to the main shrimp producers. As shrimp aquaculture has expanded and intensified, diseases have progressively become one of the most serious threats to this industry. Infectious hypodermal and hematopoietic necrosis virus (IHHNV) is an enzootic viral agent in Brazilian shrimp farms. Its is usually diagnosed by histological methods.

An optimized preparation method to obtain high-quality RNA from dry sunflower seeds

J. Yang and Ma, X. B., An optimized preparation method to obtain high-quality RNA from dry sunflower seeds, vol. 10, pp. 160-168, 2011.

In an attempt to isolate high-quality, intact total RNA from sunflower (Helianthus annuus) seeds for investigation of the molecular mechanisms of mutations, we tested various procedures, using kits, including RNAiso Plus, RNAiso Plus+RNAiso-mate for Plant Tissue, Trizol, and the Qi method, but no high-quality total RNA of high integrity was obtained with any of these methods, probably due to the high content of polyphenols, polysaccharides, and secondary metabolites in mature sunflower seeds. Modifications were made to the Qi method.

Vero cells infected with the Lederle strain of canine distemper virus have increased Fas receptor signaling expression at 15 h post-infection

H. L. Del Puerto, Martins, A. S., Braz, G. F., Alves, F., Heinemann, M. B., Rajão, D. S., Araújo, F. C., Martins, S. F., Nascimento, D. R., Leite, R. C., and Vasconcelos, A. C., Vero cells infected with the Lederle strain of canine distemper virus have increased Fas receptor signaling expression at 15 h post-infection, vol. 10, pp. 2527-2533, 2011.

We evaluated the expression of the Fas receptor gene in Vero cells infected with the Lederle vaccine strain of canine distemper virus using RT-PCR. Vero cells were plated, and after being grown for 24 h in MEM with 5% FBS, 80-90% confluent monolayer cultures were infected with the virus. The cells were harvested at 3, 6, 9, and 15 h post-infection. Uninfected Vero cells were used as a control. Total RNA was isolated from Vero cells using 1 mL Trizol® LS, and RT was performed using 2 μg total RNA.

Gene expression in swine granulosa cells and ovarian tissue during the estrous cycle

P. V. Silva, Guimarães, S. E. F., Guimarães, J. D., Neto, J. B., Lopes, P. S., Nascimento, C. Sdo, de Campos, C. F., Weller, M. M. C. A., Botelho, M. E., and Faria, V. R., Gene expression in swine granulosa cells and ovarian tissue during the estrous cycle, vol. 10, pp. 2258-2267, 2011.

The components of the insulin-like growth factor (IGF) system appear to be involved in regulation of ovarian follicular growth and atresia in the pig. We investigated the expression pattern of mRNAs for IGF1 (IGF1), its binding proteins (IGFBP1, IGFBP2, IGFBP3, and IGFBP5), and epidermal growth factor in swine follicle cells and ovarian tissue throughout the estrous cycle using the real-time quantitative PCR technique.

Molecular characterization and tissue-specific expression of invariant chain in the muscovy duck (Cairina moschata)

S. J. Liu, Chen, F. F., Dai, Y., Wu, C., Ni, Q. S., and Yu, W. Y., Molecular characterization and tissue-specific expression of invariant chain in the muscovy duck (Cairina moschata), vol. 10, pp. 2867-2880, 2011.

The invariant chain (Ii) plays an important role as a chaperone for MHC II maturation and facilitates antigen presentation in vertebrates. We cloned, characterized and made a homology analysis of healthy adult muscovy duck Ii (MDIi), from a poultry farm in the suburban district of Hefei city in China, by rapid amplification of cDNA ends (RACE)-PCR and by measuring expression of the MDIi gene in various tissues by real-time quantitative PCR. A full-length cDNA sequence of MDIi was obtained, 1188-bp long, encoding a 222-amino acid protein.

Genome-wide identification, classification, and analysis of two-component signal system genes in maize

Z. X. Chu, Ma, Q., Lin, Y. X., Tang, X. L., Zhou, Y. Q., Zhu, S. W., Fan, J., and Cheng, B. J., Genome-wide identification, classification, and analysis of two-component signal system genes in maize, vol. 10, pp. 3316-3330, 2011.

Cytokinins play many vital roles in plant development and physiology. In plants, cytokinin signals are sensed and transduced by the two-component signal system. This signaling cascade is typically composed of three proteins: a sensory histidine kinase, a histidine phosphotransfer protein, and a response regulator. Through a comprehensive genome-wide analysis of the maize (Zea mays) genome, 48 genes were identified, including 11 ZmHKs, 9 ZmHPs, and 28 ZmRRs (21 A-type ZmRRs and 7 B-type ZmRRs).

Activity and expression of polygalacturonase vary at different fruit ripening stages of sweet pepper cultivars

S. S. Ahmed, Gong, Z. - H., Khan, M. A., Yin, Y. - X., Guo, W. - L., and Imran, J., Activity and expression of polygalacturonase vary at different fruit ripening stages of sweet pepper cultivars, vol. 10, pp. 3275-3290, 2011.

Activity and expression of polygalacturonase (PG), a hydrolytic enzyme involved in ultrastructural changes in the pericarp of sweet pepper (Capsicum annaum), were investigated at different ripening stages of the pepper cultivars Mandi and Talanduo. Molecular cloning of CaPG was carried out by constructing a cDNA library from three stages of fruit ripening. Morphological determination, PG assay, RT-PCR, and ultrastructural studies were used to quantify changes in CaPG gene expression in the pericarp from green, color change and fully ripened stages.

Salt stress causes a shift in the localization pattern of germin gene expression

M. Caliskan, Salt stress causes a shift in the localization pattern of germin gene expression, vol. 8, pp. 1250-1256, 2009.

The response of plants to biotic and abiotic stress factors involves biochemical, physiological, morphological, and developmental changes. Salt stress has been the subject of extensive studies due to the low salt tolerance of many crop plants. Germin and germin-like gene products are known to be involved in various aspects of plant development, such as defense, embryonic development and response to biotic as well as abiotic stress, including salt. The responses of germin and germin-like genes to salt stress vary in different plants.

Analysis of blood processing conditions to obtain high-quality total RNA from human leukocyte concentrate

I. A. G. Pérez, Santana, S. P., Argudin, T. D., and Gardon, D. O. P., Analysis of blood processing conditions to obtain high-quality total RNA from human leukocyte concentrate, vol. 6, pp. 298-307, 2007.

Blood samples are used as a biological source to discover biomarkers of hematological and non-hematological disorders. The present study shows the impact of different experimental conditions associated with cell lysis buffer, TRI-reagent protocol and blood cell storage buffer and their correlation with the quantity, quality and Adrenomedullin gene expression levels of total RNA when RT-PCR technique is used. A leukocyte cell bank protocol is also proposed for further mRNA expression analysis using RNAlater as storage buffer.

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