Project description:MiRNA plays an important role in post-transcriptional gene regulation in plants. Whether TOR is involved in post-transcriptional gene regulation remains unclear in potato and other plants. In this study, we conducted the high-throughput sequencing of genome-wide miRNAs in the potato seedlings for profiling their expression patterns and identifying TOR related miRNAs in potato.
Project description:Purpose: MicroRNAs (miRNAs) are ubiquitous components of endogenous plant transcriptome. miRNAs are small, single-stranded and ~21 nt long RNAs which regulate gene expression at the post-transcriptional level and are known to play essential roles in various aspects of plant development and growth. Previously, a number of miRNAs have been identified in potato through in silico analysis and deep sequencing approach. However, identification of miRNAs through deep sequencing approach was limited to a few tissue types and developmental stages. This study reports the identification and characterization of potato miRNAs in three different vegetative tissues and four stages of tuber development by high throughput sequencing. Results: Small RNA libraries were constructed from leaf, stem, root and four early developmental stages of tuberization and subjected to deep sequencing, followed by bioinformatics analysis. A total of 89 conserved miRNAs (belonging to 33 families), 147 potato-specific miRNAs (with star sequence) and 112 candidate potato-specific miRNAs (without star sequence) were identified. The digital expression profiling based on TPM (Transcripts Per Million) and qRT-PCR analysis of conserved and potato-specific miRNAs revealed that some of the miRNAs showed tissue specific expression (leaf, stem and root) while a few demonstrated tuberization stage-specific expressions. Targets were predicted for identified conserved and potato-specific miRNAs, and predicted targets of four conserved miRNAs, miR160, miR164, miR172 and miR171, which are ARF16 (Auxin Response Factor 16), NAM (NO APICAL MERISTEM), RAP1 (Relative to APETALA2 1) and HAIRY MERISTEM (HAM) respectively, were experimentally validated using 5′RLM-RACE (RNA ligase mediated rapid amplification of cDNA ends). Gene ontology (GO) analysis for potato-specific miRNAs was also performed to predict their potential biological functions. Conclusions: We report a comprehensive study of potato miRNAs at genome-wide level by high-throughput sequencing and demonstrate that these miRNAs have tissue and/or developmental stage specific expression profile. Also, predicted targets of conserved miRNAs were experimentally confirmed for the first time in potato. Our findings indicate the existence of extensive and complex small RNA population in this crop and suggest their important role in pathways involved in diverse biological processes, including tuber developmental process.
Project description:Primary objectives: The primary objective is to investigate circulating tumor DNA (ctDNA) via deep sequencing for mutation detection and by whole genome sequencing for copy number analyses before start (baseline) with regorafenib and at defined time points during administration of regorafenib for treatment efficacy in colorectal cancer patients in terms of overall survival (OS).
Primary endpoints: circulating tumor DNA (ctDNA) via deep sequencing for mutation detection and by whole genome sequencing for copy number analyses before start (baseline) with regorafenib and at defined time points during administration of regorafenib for treatment efficacy in colorectal cancer patients in terms of overall survival (OS).
Project description:Nowadays, given the globalization as well as the numerous technological developments and innovations that govern the food market, consumer’s expectations, regarding the food they eat, have increased and the information they receive should be at least reliable. This proof-of- concept study is the first analysis of potato tuber tissue using integrated multi-omics approaches across genome-wide DNA methylation, RNA sequencing and quantitative proteomics to obtain the molecular portrait of famous PGI potatoes of Naxos island. Furthermore, we used metagenomics analysis in order to discriminate potato tubers produced in diverse PGI regions based on the distinct microbiological patterns. Hence, we reveal key molecular factors related to harvest and post-harvest through the dynamics of causal models.
Project description:Purpose: MicroRNAs (miRNAs) are ubiquitous components of endogenous plant transcriptome. miRNAs are small, single-stranded and ~21 nt long RNAs which regulate gene expression at the post-transcriptional level and are known to play essential roles in various aspects of plant development and growth. Previously, a number of miRNAs have been identified in potato through in silico analysis and deep sequencing approach. However, identification of miRNAs through deep sequencing approach was limited to a few tissue types and developmental stages. This study reports the identification and characterization of potato miRNAs in three different vegetative tissues and four stages of tuber development by high throughput sequencing. Results: Small RNA libraries were constructed from leaf, stem, root and four early developmental stages of tuberization and subjected to deep sequencing, followed by bioinformatics analysis. A total of 89 conserved miRNAs (belonging to 33 families), 147 potato-specific miRNAs (with star sequence) and 112 candidate potato-specific miRNAs (without star sequence) were identified. The digital expression profiling based on TPM (Transcripts Per Million) and qRT-PCR analysis of conserved and potato-specific miRNAs revealed that some of the miRNAs showed tissue specific expression (leaf, stem and root) while a few demonstrated tuberization stage-specific expressions. Targets were predicted for identified conserved and potato-specific miRNAs, and predicted targets of four conserved miRNAs, miR160, miR164, miR172 and miR171, which are ARF16 (Auxin Response Factor 16), NAM (NO APICAL MERISTEM), RAP1 (Relative to APETALA2 1) and HAIRY MERISTEM (HAM) respectively, were experimentally validated using 5M-bM-^@M-2RLM-RACE (RNA ligase mediated rapid amplification of cDNA ends). Gene ontology (GO) analysis for potato-specific miRNAs was also performed to predict their potential biological functions. Conclusions: We report a comprehensive study of potato miRNAs at genome-wide level by high-throughput sequencing and demonstrate that these miRNAs have tissue and/or developmental stage specific expression profile. Also, predicted targets of conserved miRNAs were experimentally confirmed for the first time in potato. Our findings indicate the existence of extensive and complex small RNA population in this crop and suggest their important role in pathways involved in diverse biological processes, including tuber developmental process. Total seven (Leaf, Root, Stem, Potato Tuber stage 0(PT0),Potato Tuber stage 1(PT1),Potato Tuber stage 2(PT2),Potato Tuber stage 3(PT3) ) small RNA libraries were consctructed and sequenced by deep sequencing using Illumina GAIIx.
Project description:The anthocyanin glycosides in potato tubers not only provide the plant with bright color and high nutritional value, but also have pharmacological effects such as antioxidant, hypotensive, lipid-lowering and immunity-boosting. However, the regulatory mechanism of anthocyanin in potato tubers is still unclear. In this study, twelve miRNA libraries, three of which are from the Red beauty (R-1, R-2 and R-3), three of which are from the Black kingkong (B-1, B-2 and B-3), three of which are from the Longshu No.7 (Y-1, Y-2 and Y-3) and three of which are from the LK77 (W-1, W-2 and W-3), were constructed and analyzed by high-throughput sequencing. A total of 167.3 M of clean reads were obtained. After the raw reads were subjected to quality control and filtering, 151,010,820 clean tags were obtained from the twelve libraries. The filtered reads were subsequently compared with the updated potato reference genome, and the vast majority of effective reads were successfully mapped. The mapped reads from the twelve libraries covered 77.10-84.48% of the potato genome, therefore, the analysis based on the genome was considered reliable. The lengths of the obtained miRNAs were further analysed to find that 64% of the miRNAs were concentrated at 21bp in length, followed by a higher number of miRNAs at 22bp and 24bp in length.
Project description:Plant microRNAs (miRNAs) have emerged as important regulators in developmental processes and stress responses in plants. To identify the wound-responsive miRNAs in the leaves of sweet potato, small RNA deep sequencing was conducted on unwounded and wounded leaves (30 min). Total RNAs were isolated for library construction and analyzed by RNA-sequencing via Illumina Genome Analyzer IIx platform. About 16 million total reads were obtained for each sample.
Project description:Long noncoding RNAs (lncRNAs) represent a class of RNA molecules that are implicated in regulation of gene expression, both in mammals and plants. While much progress has been made in determining the biological functions of lncRNAs in mammals, the functional roles of lncRNAs in plants are still poorly understood. Specifically, the roles of lncRNAs in plant defense responses are yet to be fully explored. Here, we used strand-specific RNA sequencing to identify 1649 lncRNAs in potato (Solanum tuberosum) from stem tissues. The lncRNAs are expressed from all 12 potato chromosomes and generally smaller in size compared to protein-coding genes. Like in other plants, most potato lncRNAs (86%) are transcribed from intergenic regions and possess single exons. A time-course RNA-seq analysis between a tolerant and susceptible potato cultivar challenged with Pectobacterium carotovorum subsp. brasilience revealed that 227 of these lncRNAs could be associated with response to this pathogen. These results suggest that lncRNAs have potential functional roles in potato defense responses. This work provides the foundation for further functional studies in understanding potato defense mechanisms.
Project description:Long noncoding RNAs (lncRNAs) represent a class of RNA molecules that are implicated in regulation of gene expression, both in mammals and plants. While much progress has been made in determining the biological functions of lncRNAs in mammals, the functional roles of lncRNAs in plants are still poorly understood. Specifically, the roles of lncRNAs in plant defense responses are yet to be fully explored. Here, we used strand-specific RNA sequencing to identify 1649 lncRNAs in potato (Solanum tuberosum) from stem tissues. The lncRNAs are expressed from all 12 potato chromosomes and generally smaller in size compared to protein-coding genes. Like in other plants, most potato lncRNAs (86%) are transcribed from intergenic regions and possess single exons. A time-course RNA-seq analysis between a tolerant and susceptible potato cultivar challenged with Pectobacterium carotovorum subsp. brasilience revealed that 227 of these lncRNAs could be associated with response to this pathogen. These results suggest that lncRNAs have potential functional roles in potato defense responses. This work provides the foundation for further functional studies in understanding potato defense mechanisms.
Project description:Background: The oomycete Phytophthora infestans possesses active RNA silencing pathways, which presumably enable this plant pathogen to control the large numbers of transposable elements present in its 240 Mb genome. Small RNAs (sRNAs), central molecules in RNA silencing, are known to also play key roles in this organism, notably in regulation of critical effector genes needed for infection of its potato host. Results: To identify additional classes of sRNAs in oomycetes, we mapped deep sequencing reads to transfer RNAs (tRNAs) thereby revealing the presence of 19-40 nt tRNA-derived RNA fragments (tRFs). Northern blot analysis identified abundant tRFs corresponding to half tRNA molecules. Some tRFs accumulated differentially during infection, as seen by examining sRNAs sequenced from P. infestans-potato interaction libraries. The putative connection between tRF biogenesis and the canonical RNA silencing pathways was investigated by employing hairpin RNA-mediated RNAi to silence the genes encoding P. infestans Argonaute (PiAgo) and Dicer (PiDcl) endoribonucleases. By sRNA sequencing we show that tRF accumulation is PiDcl1-independent, while Northern hybridizations detected reduced levels of specific tRNA-derived species in the PiAgo1 knockdown line. Conclusions: Our findings extend the sRNA diversity in oomycetes to include fragments derived from non-protein-coding RNA transcripts and identify tRFs with elevated levels during infection of potato by P. infestans.