A method for rapid isolation of total RNA of high purity and yield from <i>Arthrospira platensis</i>

Pathak, Rashmi; Lochab, Sunila

doi:10.1139/w10-045

Cited by 10 publications

(7 citation statements)

References 29 publications

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“…The seeds were vernalized prior to inoculation at 4°C for 2–3 days. Total RNA was isolated from 3–4 week old whole seedlings using a modified hot phenol-LiCl method optimized on Arabidopsis [29]and reported in the context of cyanobacteria [30]. RNA samples were analyzed by Nanodrop spectrophotometer and Bioanalyzer (Agilent technologies, Santa Clara, USA) to determine the quality, quantity and suitability for microarray.…”

Section: Methodsmentioning

confidence: 99%

Transcriptome Analysis of Arabidopsis GCR1 Mutant Reveals Its Roles in Stress, Hormones, Secondary Metabolism and Phosphate Starvation

et al. 2015

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The controversy over the existence or the need for G-protein coupled receptors (GPCRs) in plant G-protein signalling has overshadowed a more fundamental quest for the role of AtGCR1, the most studied and often considered the best candidate for GPCR in plants. Our whole transcriptome microarray analysis of the GCR1-knock-out mutant (gcr1-5) in Arabidopsis thaliana revealed 350 differentially expressed genes spanning all chromosomes. Many of them were hitherto unknown in the context of GCR1 or G-protein signalling, such as in phosphate starvation, storage compound and fatty acid biosynthesis, cell fate, etc. We also found some GCR1-responsive genes/processes that are reported to be regulated by heterotrimeric G-proteins, such as biotic and abiotic stress, hormone response and secondary metabolism. Thus, GCR1 could have G-protein-mediated as well as independent roles and regardless of whether it works as a GPCR, further analysis of the organism-wide role of GCR1 has a significance of its own.

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Section: Methodsmentioning

confidence: 99%

Transcriptome Analysis of Arabidopsis GCR1 Mutant Reveals Its Roles in Stress, Hormones, Secondary Metabolism and Phosphate Starvation

et al. 2015

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“…Total RNA was isolated by hot phenol extraction and lithium chloride precipitation method as described (Pathak and Lochab, 2010 ). Total RNA was qualitatively and quantitatively analyzed by spectrophotometry and agarose gel electrophoresis.…”

Section: Methodsmentioning

confidence: 99%

Microarray Analysis of Rice d1 (RGA1) Mutant Reveals the Potential Role of G-Protein Alpha Subunit in Regulating Multiple Abiotic Stresses Such as Drought, Salinity, Heat, and Cold

2016

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The genome-wide role of heterotrimeric G-proteins in abiotic stress response in rice has not been examined from a functional genomics perspective, despite the availability of mutants and evidences involving individual genes/processes/stresses. Our rice whole transcriptome microarray analysis (GSE 20925 at NCBI GEO) using the G-alpha subunit (RGA1) null mutant (Daikoku 1 or d1) and its corresponding wild type (Oryza sativa Japonica Nipponbare) identified 2270 unique differentially expressed genes (DEGs). Out of them, we mined for all the potentially abiotic stress-responsive genes using Gene Ontology terms, STIFDB2.0 and Rice DB. The first two approaches produced smaller subsets of the 1886 genes found at Rice DB. The GO approach revealed similar regulation of several families of stress-responsive genes in RGA1 mutant. The Genevestigator analysis of the stress-responsive subset of the RGA1-regulated genes from STIFDB revealed cold and drought-responsive clusters. Meta data analysis at Rice DB revealed large stress-response categories such as cold (878 up/810 down), drought (882 up/837 down), heat (913 up/777 down), and salt stress (889 up/841 down). One thousand four hundred ninety-eight of them are common to all the four abiotic stresses, followed by fewer genes common to smaller groups of stresses. The RGA1-regulated genes that uniquely respond to individual stresses include 111 in heat stress, eight each in cold only and drought only stresses, and two genes in salt stress only. The common DEGs (1498) belong to pathways such as the synthesis of polyamine, glycine-betaine, proline, and trehalose. Some of the common DEGs belong to abiotic stress signaling pathways such as calcium-dependent pathway, ABA independent and dependent pathway, and MAP kinase pathway in the RGA1 mutant. Gene ontology of the common stress responsive DEGs revealed 62 unique molecular functions such as transporters, enzyme regulators, transferases, hydrolases, carbon and protein metabolism, binding to nucleotides, carbohydrates, receptors and lipids, morphogenesis, flower development, and cell homeostasis. We also mined 63 miRNAs that bind to the stress responsive transcripts identified in this study, indicating their post-transcriptional regulation. Overall, these results indicate the potentially extensive role of RGA1 in the regulation of multiple abiotic stresses in rice for further validation.

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“…Total RNA was isolated from wide-type Col-0 and 2b-3 plants using hot-phenol extraction59. For 2b-associated sRNAs sequencing, aerial tissues from 6myc-SD2b transgenic plants were subjected to immunoprecipitation using anti-α-myc Agarose (Abmart).…”

Section: Methodsmentioning

confidence: 99%

Genome-wide identification of endogenous RNA-directed DNA methylation loci associated with abundant 21-nucleotide siRNAs in Arabidopsis

Zhao

Fang

Duan

et al. 2016

Sci Rep

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In Arabidopsis, the 24-nucleotide (nt) small interfering RNAs (siRNAs) mediates RNA-directed DNA methylation (RdDM) and transcriptional gene silencing (TGS) of transposable elements (TEs). In the present study, we examined genome-wide changes in DNA methylation and siRNA accumulation in Arabidopsis induced by expression of the Cucumber mosaic virus silencing suppressor protein 2b known to directly bind to both the 21/24-nt siRNAs as well as their associated Argonaute proteins. We demonstrated a genome-wide reduction of CHH and CHG methylation in the 2b-transgenic plants. We found that 2b suppressed RdDM not only at the previously annotated loci directed by 24-nt siRNAs but also a new set of loci associated with 21/22-nt siRNAs. Further analysis showed that the reduced methylation of TEs and coding genes targeted by 21/22-nt siRNAs was associated with sequestration of the duplex siRNAs by the 2b protein but not with changes in either siRNA production or transcription. Notably, we detected both the deletion and/or the transposition of multicopy TEs associated with 2b-induced hypomethylation, suggesting potential TE reactivation. We propose that the silencing of many TEs in Arabidopsis is controlled by the 24- and 21-nt endogenous siRNAs analogous to Drosophila TE silencing by PIWI-interacting RNAs and siRNAs.

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A method for rapid isolation of total RNA of high purity and yield from Arthrospira platensis

Cited by 10 publications

References 29 publications

Transcriptome Analysis of Arabidopsis GCR1 Mutant Reveals Its Roles in Stress, Hormones, Secondary Metabolism and Phosphate Starvation

Transcriptome Analysis of Arabidopsis GCR1 Mutant Reveals Its Roles in Stress, Hormones, Secondary Metabolism and Phosphate Starvation

Microarray Analysis of Rice d1 (RGA1) Mutant Reveals the Potential Role of G-Protein Alpha Subunit in Regulating Multiple Abiotic Stresses Such as Drought, Salinity, Heat, and Cold

Genome-wide identification of endogenous RNA-directed DNA methylation loci associated with abundant 21-nucleotide siRNAs in Arabidopsis

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