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PMID: 16668285;PMCID: PMC1080875;Abstract:
Microfluorometric analysis of the nuclear DNA contents of the somatic tissues of Arabidopsis thaliana has revealed extensive endoreduplication, resulting in tissues that comprise mixtures of polyploid cells. Endoreduplication was found in all tissues except those of the inflorescences and was developmentally regulated according to the age of the tissues and their position within the plant.
Postier, B. L., Wang, H., Singh, A., Impson, L., Andrews, H. L., Klahn, J., Hong, L. i., Risinger, G., Pesta, D., Deyholos, M., Galbraith, D. W., Sherman, L. A., & Burnap, R. L. (2003). The construction and use of bacterial DNA microarrays based on an optimized two-stage PCR strategy. BMC Genomics, 4.
Abstract:
Background: DNA microarrays are a powerful tool with important applications such as global gene expression profiling. Construction of bacterial DNA microarrays from genomic sequence data using a two-stage PCR amplification approach for the production of arrayed DNA is attractive because it allows, in principal, the continued re-amplification of DNA fragments and facilitates further utilization of the DNA fragments for additional uses (e.g. over-expression of protein). We describe the successful construction and use of DNA microarrays by the two-stage amplification approach and discuss the technical challenges that were met and resolved during the project. Results: Chimeric primers that contained both gene-specific and shared, universal sequence allowed the two-stage amplification of the 3,168 genes identified on the genome of Synechocystis sp. PCC6803, an important prokaryotic model organism for the study of oxygenic photosynthesis. The gene-specific component of the primer was of variable length to maintain uniform annealing temperatures during the 1st round of PCR synthesis, and situated to preserve full-length ORFs. Genes were truncated at 2 kb for efficient amplification, so that about 92% of the PCR fragments were full-length genes. The two-stage amplification had the additional advantage of normalizing the yield of PCR products and this improved the uniformity of DNA features robotically deposited onto the microarray surface. We also describe the techniques utilized to optimize hybridization conditions and signal-to-noise ratio of the transcription profile. The inter-lab transportability was demonstrated by the virtual error-free amplification of the entire genome complement of 3,168 genes using the universal primers in partner labs. The printed slides have been successfully used to identify differentially expressed genes in response to a number of environmental conditions, including salt stress. Conclusions: The technique detailed here minimizes the cost and effort to replicate a PCRgenerated DNA gene fragment library and facilitates several downstream processes (e.g. directional cloning of fragments and gene expression as affinity-tagged fusion proteins) beyond the primary objective of producing DNA microarrays for global gene expression profiling. © 2003 Postier et al; licensee BioMed Central Ltd.
Baisakh, N., Ramanarao, M. V., Rajasekaran, K., Subudhi, P., Janda, J., Galbraith, D., Vanier, C., & Pereira, A. (2012). Enhanced salt stress tolerance of rice plants expressing a vacuolar H +-ATPase subunit c1 (SaVHAc1) gene from the halophyte grass Spartina alterniflora Löisel. Plant Biotechnology Journal, 10(4), 453-464.
PMID: 22284568;Abstract:
The physiological role of a vacuolar ATPase subunit c1 (SaVHAc1) from a halophyte grass Spartina alterniflora was studied through its expression in rice. The SaVHAc1-expressing plants showed enhanced tolerance to salt stress than the wild-type plants, mainly through adjustments in early stage and preparatory physiological responses. In addition to the increased accumulation of its own transcript, SaVHAc1 expression led to increased accumulation of messages of other native genes in rice, especially those involved in cation transport and ABA signalling. The SaVHAc1-expressing plants maintained higher relative water content under salt stress through early stage closure of the leaf stoma and reduced stomata density. The increased K +/Na + ratio and other cations established an ion homoeostasis in SaVHAc1-expressing plants to protect the cytosol from toxic Na + and thereby maintained higher chlorophyll retention than the WT plants under salt stress. Besides, the role of SaVHAc1 in cell wall expansion and maintenance of net photosynthesis was implicated by comparatively higher root and leaf growth and yield of rice expressing SaVHAc1 over WT under salt stress. The study indicated that the genes contributing toward natural variation in grass halophytes could be effectively manipulated for improving salt tolerance of field crops within related taxa. © 2012 Louisiana State University Agricultural Center. Plant Biotechnology Journal © 2012 Society for Experimental Biology, Association of Applied Biologists and Blackwell Publishing Ltd.
