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Transcriptomic analysis of Grapevine (cv. Summer Black) leaf at four developmental stages, using the Illumina Platform

ABSTRACT: conducted to find out the transcriptional system that is connected with the development of leaves and production of metabolites during photosynthesis. We examine the cellular expression by use the up-to-date RNA sequencing expertise to carry out a complete analysis of the transcriptional profile of V. vinifera leaves at four different developmental stages.

ORGANISM(S): Vitis vinifera

PROVIDER: GSE74428 | GEO | 2015/10/29

SECONDARY ACCESSION(S): PRJNA300400

REPOSITORIES: GEO

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Project description:Proceeding to illumina sequencing, determining RNA integrity numbers for poly RNA were separated from each of the four developmental stages of cv. Summer Black leaves by using Illumina HiSeq™ 2000. The sums of 272,941,656 reads were generated from vitis vinifera leaf at four different developmental stages, with more than 27 billion nucleotides of the sequence data. At each growth stage, RNA samples were indexed through unique nucleic acid identifiers and sequenced. KEGG annotation results depicted that the highest number of transcripts in 2,963 (2Avs4A) followed by 1Avs4A (2,920), and 3Avs4A (2,294) out of 15,614 (71%) transcripts were recorded. In comparison, a total of 1,532 transcripts were annotated in GOs, including Cellular component, with the highest number in "Cell part" 251 out of 353 transcripts (71.1%), followed by intracellular organelle 163 out of 353 transcripts (46.2%), while in molecular function and metabolic process 375 out of 525 (71.4%) transcripts, multicellular organism process 40 out of 525 (7.6%) transcripts in biological process were most common in 1Avs2A. While in case of 1Avs3A, cell part 476 out of 662 transcripts (71.9%), and membrane-bounded organelle 263 out of 662 transcripts (39.7%) were recorded in Cellular component. In the grapevine transcriptome, during the initial stages of leaf development 1Avs2A showed single transcript was down-regulated and none of them were up-regulated. While in comparison of 1A to 3A showed one up-regulated (photosystem II reaction center protein C) and one down regulated (conserved gene of unknown function) transcripts, during the hormone regulating pathway namely SAUR-like auxin-responsive protein family having 2 up-regulated and 7 down-regulated transcripts, phytochrome-associated protein showed 1 up-regulated and 9 down-regulated transcripts, whereas genes associated with the Leucine-rich repeat protein kinase family protein showed 7 up-regulated and 1 down-regulated transcript, meanwhile Auxin Resistant 2 has single up-regulated transcript in second developmental stage, although 3 were down-regulated at lateral growth stages (3A and 4A). In the present study, 489 secondary metabolic pathways related genes were identified during leaf growth, which mainly includes alkaloid (40), anthocyanins (21), Diterpenoid (144), Monoterpenoid (90) and Flavonoids (93). Quantitative real-time PCR was applied to validate 10 differentially expressed transcripts patterns from flower, leaf and fruit metabolic pathways at different growth stages.

Project description:Infection by the pathogen grape powdery mildew (Erysiphe necator) causes changes in the transcriptome of its susceptible host Vitis vinifera. Infection triggers the host to synthesize the signaling molecule salicylic acid (SA) which regulates the expression of a broad range of defense-related plant genes. In addition, it is hypothesized that E. necator directly modulates gene expression in V. vinifera via the haustorial complex. This microarray experiment was designed to dissect host transcriptome changes triggered directly by E. necator infection and indirectly through the SA response. We accomplished this by conducting two separate global leaf transcriptome analyses using the Vitis Affymetrix GeneChip platform: in one, we compared the leaves with fully established PM colonies to healthy reference leaves, in another, we compared healthy leaves with artificially elevated SA levels to healthy reference leaves. Overlaying host transcriptome changes from these two experiments enabled us to glean out V. vinifera genes that modulate their expression in response E. necator in an SA-independent manner. We characterized gene expression profiles in (1) healthy reference leaves, (2) leaves with well-established by E. necator colonies, and (3) healthy leaves in which SA levels were artificially elevated by the presence of methyl salicylate (15 µM) in the atmosphere. Treatments (1) and (2) differed only in the presence E. necator infection, whereas treatments (1) and (3) differed only in SA levels. Consequently, statistical comparisons were made only between signal intensities from treatments (1) and (2) and from treatments (1) and (3). Each treatment was done in three biological repeats, that is, each experiment was repeated three times in 14-day intervals with dedicated biological material (plants were used in a single replicate and not reused in another repeat or treatment). Each biological replicate consisted of 10 potted vines. For RNA extraction, two young leaves were harvested from each of the 10 vines and pooled into a single sample.

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Transcriptomic analysis of Grapevine (cv. Summer Black) leaf at four developmental stages, using the Illumina Platform

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