Project description:We have developed a serum-free chemical defined medium, namely 6C, that can directly convert mouse embryonic fibroblast (MEFs), glia cells into neurons in vitro. Human cells such as human foreskin fibroblast(HHFs), Hela cells and born marrow derived mesenchymal stem cells (BM-hMSC) can also be converted into neuron-like cells by this medium with some modification such as including several other small molecules. To understand the possible mechanisms, the transdifferentiation of MEFs by 6C was chosen as a model system and gene profiling at different time point during the conversion was carried out by RNA sequencing using Illumina MiSeq. MEFs was maintained in MEF medium (DMEM containing 10% FBS, 1mM Glutamax and 100X NEAA), to start the induction, the culture medium was shift to 6C and marked as day 0, neurons could be generated in day 15. mRNA samples was collected at day 0, 2, 5, 10, 15 during the process, with cell lysed by Trizol and mRNA enriched by Illumina TruSeq RNA Sample Preparation v2 kit. We find that most cell cycle related genes were up regulated during the first few days of induction, while many Notch pathway genes up regulated during the later phase of this process. The RNA sequencing data provided important cues for further study on the mechanisms of the direct neuronal induction process. Gene profiling at different time point during the transdifferentiation mediated by 6C medium.
Project description:TNF is a proinflammatory cytokine with established roles in host defense and immune system organogenesis. Here we report a novel physiological function of TNF that extends its effect beyond the host into the developing offspring. A partial/complete maternal TNF-deficit, specifically in hematopoietic cells, resulted in reduced milk levels of chemokines IP-10, MCP-1/-3/-5, and MIP-1β, which in turn, augmented offspring postnatal hippocampal proliferation, leading to improved adult spatial memory. These effects were reproduced by the postpartum administration of a clinically used anti-TNF agent. Chemokines, fed to suckling pups of TNF-deficient mothers, restored both postnatal proliferation and adult spatial memory to normal levels. This work identifies a TNF-dependent “lactrocrine” pathway that programs offspring hippocampal development and memory. The level of ambient TNF is known to be downregulated by physical activity/exercise and adaptive stress; thus, we propose that the maternal TNF-milk chemokine pathway evolved to promote offspring adaptation to post-weaning environmental challenges/competition. Examined transcriptomes of TNF wild type offspring of TNF wild type or heterozygouse mothers
Project description:Next Generation Sequencing with differdent pluripotent transcript factor overexpression in MEF Somatic cells can be reprogrammed into induced pluripotent stem cells (iPSCs) by defined factors. The low efficiency of reprogramming limited the potential application of iPSCs. Here we found that knockdown LSD1 , which demethylates histone H3 Lys 4 or 9 , could increase iPSCs generation. It has been reported that LSD1 interaction with Oct4 which is the core factor of reprogramming. So we try to find out the different of overexpression Oct4 or Kllf4/Sox2 in MEF and LSD1 inhibitor. Retrovirus-mediated different pluripotent transcript factor overexpression in MEF cells after 4 days collect mRNA profiles processing with Illumina MiSeq; MEF, Mouse Embryonic Fibroblast O, MEF infected with Oct4 KS,MEF infected with Klf4 and Sox2 OSK, MEF infected with Oct4 Klf4 and Sox2 T20, MEF infected with Oct4 Klf4 and Sox2 and treated with 20nm LSD1 inhibitor Tranylcypromine
Project description:Purpose: The goals of this study are using RNA-seq to obtain cucumber and Botrytis cinerea transcriptome changes during infection Methods: mRNA profiles of anti-infection samples and interaction sample were generate by deep sequencing,using Illumina Hiseq 2500. The sequence reads that passed quality filters were analyzed at the transcript isoform level with two methods: BurrowsâWheeler Aligner (BWA) followed by ANOVA (ANOVA) and TopHat followed by Cufflinks. qRTâPCR validation was performed using SYBR Green assays Results: Using an optimized data analysis workflow,In total, 248,908,688 raw reads were generated; after removing low-quality reads and those containing adapter and poly-N, 238,341,648 clean reads remained to map the reference genome. There were 3,512 cucumber (differential expression genes) DEGs and 1,735 B. cinerea DEGs. GO enrichment and KEGG enrichment analysis were performed on these DEGs to study the interaction between cucumber and B. cinerea. To verify the reliability and accuracy of our transcriptome data, 5 cucumber DEGs and 5 B. cinerea DEGs were chosen for RT-PCR verification. Conclusions:To the best of our knowledge, this is the first analysis of large-scale transcriptome changes of cucumber during the infection of Botrytis cinerea. These results will increase our understanding of the molecular mechanisms of the cucumber defense Botrytis cinerea and may be used to protect plants against disasters caused by necrotrophic fungal pathogens. mRNA profiles of infection and anti-infection cucumber were generated by deep sequencing, using Illumina Hiseq 2500 .
Project description:Purpose: We report the application of NGS for profiling the impacts of BDE47 exposure on the transcriptome of zebrafish larvae. Methods: mRNA profiles of 6-day-old BDE47-treated and control zebrafish larvae were generated by deep sequencing using Illumina Hisq 2000 platform. The sequence reads that passed quality filters were analyzed at the transcript isoform level with TopHat followed by Cufflinks. Results: Compared BDE47 treatments with solvent control, 2235 transcripts were affected after 500 μg/l exposure, in which 1338 were up-regulated and 897 were down-regulated, and 552 transcripts were affected after 5 μg/l exposure, in which 155 were up-regulated and 397 were down-regulated. High concentration of BDE47 exposure resulted in more up-regulated genes. Conclusions: This study provides a framework for the application of high-throughput RNA sequencing towards characterization of the impacts of BDE47 on whole zebrafish larval transcriptome. Examination of zebrafish larvae transcriptomes with vehicle and 2 different concentrations of BDE47 treatments.
Project description:We report the application of single-molecule-based sequencing technology for high-throughput profiling of different ovine muscle's transcriptomes. 9.27 gigabases of sequence from two different breeds of sheep. Examination of 2 different muscle's transcriptomes in 2 breed types.
Project description:Understanding how genomes encode complex cellular and organismal behaviors is an outstanding challenge of modern genetics. Unlike classical screening methods, analysis of the genetic variation that occurs naturally in wild populations can enable rapid, genome-scale trait mapping with a medium-throughput experimental design. Here we describe the results of the first genome-wide association analysis (GWAS) of a microbial eukaryote, using wild isolates of the filamentous fungus Neurospora crassa. We transcriptionally profiled each of 112 individuals collected in Louisiana and used RNA-seq data to genotype each strain at thousands of genetic loci genome-wide. We used these genotypes in a mapping analysis of microbial communication. In N. crassa, germinated asexual spores (germlings) sense the presence of other germlings, grow toward them in a coordinated fashion, and fuse. We evaluated germlings of each strain for their ability to chemically sense, chemotropically seek, and undergo cell fusion. GWAS identified one gene, NCU04379 (cse-1, encoding a homolog of neuronal calcium sensor), at which inheritance was strongly associated with the efficiency of germling communication. Deletion of cse-1 significantly lowered germling communication and fusion, and two genes encoding predicted binding partners of CSE-1 were also required for the communication trait. Additionally, mining our association results for signaling and secretion genes with a potential role in germling communication, we validated six more novel determinants of germling communication, including a secreted protease and two other genes whose deletion conferred a novel phenotype of increased communication and multi-germling fusion. These results establish protein secretion as a linchpin of germling communication in N. crassa and shed light on the regulation of communication molecules in this fungus. Our study demonstrates the power of population-genetic analyses for the rapid identification of genes contributing to complex traits in microbial species. 112 mycelial mRNA profiles of 16 hour wild type (WT) Neurospora crassa strains were generated by deep sequencing, using Illumina GAIIx.
Project description:We explored the transcriptomic changes of synthetic Brassica allohexaploid by comparing to its parents using a high-throughput RNA-Seq method. A total of 35644409 sequence reads were generated, and 32642 genes were aligned from the data. There were 29260, 29060 and 29697 genes identified in Brassica rapa, Brassica carinata, and Brassica allohexaploid, respectively. We screened differentially expressed genes (DEGs) by a standard of two-fold or greater change in expression and false discovery rate (FDR) no more than 0.001. As a result, 7397 DEGs were detected between Brassica hexaploid and its parents. A large proportion of the 3184 DEGs between Brassica hexaploid and its paternal parent B. rapa was involved in biosynthesis of secondary metabolites, plant-pathogen interaction, photosynthesis, and circadian rhythm. Between Brassica hexaploid and its maternal parent B. carinata, 2233 DEGs were screened. A lot of them had functions of plant-pathogen interaction, plant hormone signal transduction, ribosome, limonene and pinene degradation, photosynthesis, and also biosynthesis of secondary metabolites. In addition, we found many transcription factor genes, methyltransferase and methylation genes that showed differential expression between Brassica hexaploid and its parents. Leaf mRNA profiles of Brassica rapa, Brassica carinata, and Brassica allohexaploid
Project description:Nucleosome organization determines chromatin state, which subsequently controls genes expression or silencing. Nucleosome remodeling occurs during somatic cell reprogramming, but it remains undetermined to what degree the re-established nucleosome organization resembles between induced pluripotent stem cells (iPSCs) and embryonic stem cells (ESCs). Here, we generated genome-wide maps of nucleosomes in mouse ESCs and iPSCs reprogrammed from somatic cells belonging to three different germ layers using a secondary reprogramming system. Pairwise comparisons show that nucleosome organization is nearly identical between ESCs and iPSCs regardless of the iPSCs’ tissue of origin. A distinct nucleosome occupancy pattern was observed at silent transcriptional units. Transcription factor binding sites possess characteristic nucleosomal architecture such that their access is governed under rotational setting and translational setting accordingly. Gene expression profiles further reveal that the transcriptional programs are highly correlated between iPSCs and ESCs. These findings indicate that nucleosome organizations can be accurately remodeled during nuclear reprogramming. Gene expression profiles of 5 cell lines with or without biological replicates
Project description:A colony of fungus is comprised of long, branching filamentous cells called hyphae. Genetic mechanisms underlying the development of hyphae are poorly understood. We sectioned hyphae of a model fungus, Neurospora crassa (pink bread mold), into six parts depending on the age of the cells; 1 hour, 3 hour, 9 hour, 15 hour, 21 hour and 27 hour old, respectively. This data submission reflects an RNAseq analysis of mRNA extracted from the 1 hour time-point. 1 mycelial mRNA profile of 1 hour growth (hyphal tip) from Neurospora crassa strain FGSC 2489, Data was generated using Illumina GAIIx.