The aim of this study is to identify Arabidopsis genes whose expression is altered by aphid feeding. An understanding of the plant aphid interaction at the level of the plant transcriptome will 1) consolidate current areas of investigation focused on the phloem composition (the aphid diet), 2) open up areas of plant aphid interactions for ourselves and other workers, 3) Contribute to understanding the use of new molecular technologies in an environmental context and 4) contribute to existing and development of novel control strategies.Our Arabidopsis/Myzus persicae system provides a valuable model for the study because of: a) the advantages of using Arabidopsis, b) The ability to use clonal insects, c) phloem feeding aphids facilitate focus on a specific cell type, d) aphid stylectomy allows collection of pure phloem sap to monitor phloem phenotype of the plant and the insect diet, e) we have techniques to monitor the reproductive performance and feeding behaviour aphids.Our strategy has been to test the function of selected genes, particularly those regulating phloem composition (the feeding site of the aphid) based on current phloem models of phloem function. Gene choice is limited the simplicity of current models of phloem aphid interaction.We propose a simple two treatment (aphid infested vs control plants) experiment that will identify novel target genes for future analysis. Arabidopsis plants (variety Columbia) will be grown in 16/8 light/dark in temperature controlled growth rooms. At growth stage 3.90, when rosette growth is complete, 10 clonal adult Myzus persicae will be caged in clip cages on the two largest leaves on each plant. Control plants will be treated identically except that the cages will be empty. Leaves will be harvested 8 h after infestation. This time point is selected as we know that 90% of aphids are plugged into the sieve element within 2h and that a 6h lag phase has period has previously been used when examining gene expression affected by wounding. In subsequent experiments we will examine time courses of expression of relevant genes using other approaches. Pooling two leaves from each of ten plants will generate the RNA sample, ensuring that expression signals are representative of the population of plants.
Exploring plant responses to aphid feeding using a full Arabidopsis microarray reveals a small number of genes with significantly altered expression.
Specimen part
View SamplesAbstract Background Traumatic brain injury (TBI) results in irreversible damage at the site of impact and initiates cellular and molecular processes that lead to secondary neural injury in the surrounding tissue. We used microarray analysis to determine which genes, pathways and networks were significantly altered using a rat model of TBI. Adult rats received a unilateral controlled cortical impact (CCI) and were sacrificed 24h post-injury. The ipsilateral hemi-brain tissue at the site of the injury, the corresponding contralateral hemi-brain tissue, and nave (control) brain tissue were used for microarray analysis. Ingenuity Pathway Analysis (IPA) software was used to identify molecular pathways and networks that were associated with the altered gene expression in brain tissues following TBI. Results Inspection of the top fifteen biological functions in IPA associated with TBI in the ipsilateral tissues revealed that all had an inflammatory component. IPA analysis also indicated that inflammatory genes were altered on the contralateral side, but many of the genes were inversely expressed compared to the ipsilateral side. The contralateral gene expression pattern suggests a remote anti-inflammatory molecular response. We created a network of the inversely expressed common (i.e., same gene changed on both sides of the brain) inflammatory response (IR) genes and those IR genes included in pathways and networks identified by IPA that changed on only one side. We ranked the genes by the number of direct connections each had in the network, creating a gene interaction hierarchy (GIH). Two well characterized signaling pathways, toll-like receptor/NF-kappaB signaling and JAK/STAT signaling, were prominent in our GIH. Conclusions Bioinformatic analysis of microarray data following TBI identified key molecular pathways and networks associated with neural injury following TBI. The GIH created here provides a starting point for investigating therapeutic targets in a ranked order that is somewhat different than what has been presented previously. In addition to being a vehicle for identifying potential targets for post-TBI therapeutic strategies, our findings can also provide a context for evaluating the potential of therapeutic agents currently in development.
Gene expression patterns following unilateral traumatic brain injury reveals a local pro-inflammatory and remote anti-inflammatory response.
Specimen part, Treatment
View SamplesTemporal changes in the embryo transcriptome between the blastocyst stage (Day 7) and initiation of elongation (Day 13) differ between in vivo- and in vitro-derived embryos and are reflective of subsequent developmental fate.
Transcriptome changes at the initiation of elongation in the bovine conceptus.
Specimen part
View SamplesThe aim of this study was to compare the transcriptome of the different regions of the oviduct between pregnant and cyclic heifers. After synchronizing crossbred beef heifers, those in standing oestrus (=Day 0) were randomly assigned to cyclic (non bred, n=6), or pregnant (artificially inseminated, n=11) groups. They were slaughtered on Day 3 and both oviducts from each animal were isolated and cut in half to separate ampulla and isthmus. Each portion was flushed to confirm the presence of an oocyte/embryo and was then opened longitudinally and scraped to obtain epithelial cells which were snap-frozen. Oocytes and embryos were located in the isthmus of the oviduct ipsilateral to the corpus luteum. Microarray analysis of oviductal cells revealed that proximity to the corpus luteum did not affect the transcriptome of the isthmus, irrespective of pregnancy status. However, 2287 genes were differentially expressed (P<0.01) between the ampulla and isthmus of the oviduct ipsilateral to the corpus luteum. Gene ontology revealed that the main biological processes overrepresented in the isthmus were synthesis of nitrogen, lipids, nucleotides, steroids and cholesterol as well as vesicle-mediated transport, cell cycle, apoptosis, endocytosis and exocytosis, whereas cell motion, motility and migration, DNA repair, calcium ion homeostasis, carbohydrate biosynthesis and regulation of cilium movement and beat frequency were overrepresented in the ampulla. In conclusion, large differences in gene expression were observed between the isthmus and ampulla that reflect morphological and functional characteristics of each segment.
Spatial differences in gene expression in the bovine oviduct.
Specimen part
View SamplesThe objective of this study was to examine the effect of the presence of a single or multiple embryo(s) on the transcriptome of the bovine oviduct. In Experiment 1, cyclic (non-bred, n = 6) and pregnant (artificially inseminated, n = 11) heifers were slaughtered on Day 3 after estrus, and the ampulla and isthmic regions of the oviduct ipsilateral to the corpus luteum were separately flushed. Oviductal epithelial cells from the isthmus region, in which all oocytes/embryos were located, were snap-frozen for microarray analysis. In Experiment 2, heifers were divided into cyclic (non-bred, n = 6) or pregnant (multiple embryo transfer, n = 10) groups. In vitro-produced presumptive zygotes were transferred endoscopically to the ipsilateral oviduct on Day 1.5 post estrus (n = 50 zygotes per heifer). Heifers were slaughtered on Day 3 and oviductal isthmus epithelial cells were recovered for RNA sequencing. Microarray analysis in Experiment 1 failed to detect any difference in the transcriptome of the oviductal isthmus induced by the presence of a single embryo. In Experiment 2, following multiple embryo transfer, RNA sequencing revealed 278 differentially expressed genes of which 123 were up- and 155 were down-regulated in pregnant heifers. Most of the down-regulated genes were related to immune function. Overall design: Transcriptional profiles of oviductal isthmus epithelial cells from cyclic and pregnant heifers were generated by sequencing of total RNA on the Illumina HiSeq 2500 platform
Oviduct-Embryo Interactions in Cattle: Two-Way Traffic or a One-Way Street?
Specimen part, Treatment, Subject
View SamplesThis SuperSeries is composed of the SubSeries listed below.
Oviduct-Embryo Interactions in Cattle: Two-Way Traffic or a One-Way Street?
Specimen part, Treatment
View SamplesThe objective of this study was to examine the effect of the presence of a single or multiple embryo(s) on the transcriptome of the bovine oviduct. In Experiment 1, cyclic (non-bred, n = 6) and pregnant (artificially inseminated, n = 11) heifers were slaughtered on Day 3 after estrus, and the ampulla and isthmic regions of the oviduct ipsilateral to the corpus luteum were separately flushed. Oviductal epithelial cells from the isthmus region, in which all oocytes/embryos were located, were snap-frozen for microarray analysis. In Experiment 2, heifers were divided into cyclic (non-bred, n = 6) or pregnant (multiple embryo transfer, n = 10) groups. In vitro-produced presumptive zygotes were transferred endoscopically to the ipsilateral oviduct on Day 1.5 post estrus (n = 50 zygotes per heifer). Heifers were slaughtered on Day 3 and oviductal isthmus epithelial cells were recovered for RNA sequencing. Microarray analysis in Experiment 1 failed to detect any difference in the transcriptome of the oviductal isthmus induced by the presence of a single embryo. In Experiment 2, following multiple embryo transfer, RNA sequencing revealed 278 differentially expressed genes of which 123 were up- and 155 were down-regulated in pregnant heifers. Most of the down-regulated genes were related to immune function.
Oviduct-Embryo Interactions in Cattle: Two-Way Traffic or a One-Way Street?
Specimen part
View SamplesThis study relates to embryo-maternal interaction. The aim was to compare the transcriptome and ability of the ipsilateral and contralateral uterine horns to support preimplantation conceptus survival and growth to Day 14. Although differences in gene expression exist between the endometrium of uterine horns ipsilateral and contralateral to the CL in cattle, they do not impact conceptus survival or length between Days 7 and 14. Overall design: The endometrial samples from both uterine horns were collected from synchronized heifers slaughtered on Day 5, 7, 13 or 16 post-estrus (n = 5 per time) and subjected to RNA sequencing.
Do differences in the endometrial transcriptome between uterine horns ipsilateral and contralateral to the corpus luteum influence conceptus growth to day 14 in cattle?
Specimen part, Subject, Time
View SamplesAn increase in circulating progesterone (P4) concentrations is associated with increased pregnancy success in beef and dairy cattle. Our objective was to ascertain differential effects of elevated P4 concentrations following conception on endometrial gene expression in beef heifers on Days 5, 7, 13 and 16 of pregnancy, corresponding to the morula, blastocyst, elongation and maternal recognition of pregnancy stages, respectively. Estrus was synchronized in beef heifers (N=263). Two-thirds (N=140) were inseminated (Day 0), and all animals were randomly assigned to one of the following treatments: (i) pregnant, high P4; (ii) pregnant, normal P4; (iii) cycling, high P4; (iv) and cycling, normal P4. All high P4 groups received a P4 release intravaginal device (PRID) on Day 3 post-estrus/mating. Tissue was collected on Days 5, 7, 13 or 16 of the cycle or pregnancy, and pregnancy was confirmed by the presence of an appropriately developed embryo/conceptus. PRID insertion elevated (P<0.05) P4 concentrations from Day 3.5 to 8 compared with untreated animals and conceptus size was larger (P<0.05) in animals with elevated P4 on Days 13 and 16 compared with normal P4. Total RNA was extracted from predominantly intercaruncular endometria from the ipsilateral uterine horn. Samples from individual heifers were selected on the basis of their P4 profiles and gene expression was analyzed using bovine Affymetrix microarrays (N=5 per treatment per time point). Microarray data from analyses using Bioconductor GCRMA and Limma packages were subjected to a modified t-test and P-values were adjusted for multiple testing using the Benjamin and Hochberg false discovery rate method. Differentially expressed genes were selected on the basis of an adjusted P-value of <0.01. There were no detectable differences in gene expression in endometria from pregnant and cyclic heifers on Days 5, 7 and 13 post-estrus, but, the expression of 764 genes was altered due to the presence of the conceptus at maternal recognition of pregnancy (Day 16). On Days 5 and 7, elevated P4 in pregnant heifers, altered the expression of 36 and 124 genes respectively but on Days 13 and 16 there were relatively few DEG between high and normal P4 heifers (15 and 25). Of the genes that were differentially regulated by P4, the majority were unique to a specific day of the estrous cycle/early pregnancy. In conclusion, gene expression in endometria did not differ between pregnant and cycling heifers until Day 16 of pregnancy (i.e. the time of maternal recognition of pregnancy and production of interferon tau by conceptus trophectoderm); however, elevating P4 in early pregnancy programmed changes in gene expression in endometria that are hypothesized to impact early conceptus growth and development. Thus, on Days 5, 7 and 13 differential gene expression was affected by P4, but on Day 16 the conceptus primarily influenced gene expression in uterine endometria of heifers.
Conceptus-induced changes in the endometrial transcriptome: how soon does the cow know she is pregnant?
Specimen part, Time
View SamplesCellular mechanisms that contribute to low estradiol concentrations produced by the preovulatory ovarian follicle in cattle with a compromised metabolic status (such as lactatino) are largely unknown. To gain insight into the main metabolic mechanisms affecting preovulatory follicle function RNAseq profiling was conducted on non-lactating Holstein-Friesian heifers (n=16) and lactating Holstein-Friesian cows (n=17) at three stages of preovulatory follicle development: A) newly selected dominant follicle in the luteal phase (Selection); B) follicular phase before the LH surge (Differentiation) and C) pre-ovulatory phase after the LH surge (Luteinization). Based on a combination of RNA sequencing, ingenuity pathway analysis and Q-RT-PCR validation several important molecular markers involved in steroid biosynthesis, such as the expression of steroidogenic acute regulatory protein (STAR) within developing dominant follicles, were identified to be affected (downregulated) by the catabolic state. We propose that the adverse metabolic environment caused by lactation decreases preovulatory follicle function by affecting cholesterol transport into the mitochondria to initiate steroidogenesis. Overall design: Granulosa and Theca samples from the dominant follicle were taken from cows and heifers at stages: selection, differentiation and luteinization.
Effect of the metabolic environment at key stages of follicle development in cattle: focus on steroid biosynthesis.
Specimen part, Subject
View Samples