3D-RNA-seq Analysis Tool

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Published on: November 15, 2019

At the end of October GARNet hosted a workshop on the the 3d RNAseq App that has been developed by Wenbin Guo and Runxuan Zhang at the James Hutton Institute. Workshop schedule and attendees can be downloaded here (PDF).

The 3D RNA-Seq Analysis Tool is designed for the comprehensive differential expression, alternative splicing analysis and visualisation of RNASeq Data.

https://3drnaseq.hutton.ac.uk/app_direct/3DRNAseq/

It runs the analysis through a user-friendly graphical interface, that can handle complex experimental designs, allows user setting of statistical parameters, visualizes the results through graphics and tables, and generates publication quality figures and customerised  analysis reports.


The program is designed to be run by biologists with minimal bioinformatics experience (or by bioinformaticians) allowing lab scientists to take control of the analysis of their RNA-seq data.

The designers also provide a explanation on how RNAseq FastQ reads can be processed through Salmon or Kallisto using the Galaxy platform.

This entire process allows non-experts to take their raw data through to detailed outputs within a couple of days!


Instructional documents can be downloaded here and please check out the accompanying YouTube video.

  1. Transcript quantification using Galaxy (HTML)
  2. 3D RNAseq Manual (zipped HTML)
  3. 3D Workshop presentation (PPT)

CONNECTED Vector-borne Plant Virus Training Program

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Published on: November 7, 2019

A group of African early career researchers feature in a new short film about vector-borne plant viruses that devastate crops in Sub-Saharan African countries.

The seven-minute film focuses on the ground-breaking Virus-Vector Vice-Versa training course for early career researchers, run by the CONNECTED Network, which took place at The University of Bristol, UK, in summer 2019.


A group of early career researchers from 11 African countries took up fully-funded places at the course. The film features interviews with delegates discussing: 

  • what they learned on the course
  • how they will use their new skills and knowledge, and
  • the value to them of the collaborative CONNECTED network.

There are also clips from interviews with:

  • Prof. Nicola Spence, Defra, UK (Chair of the CONNECTED Management Board)
  • Prof. Gary Foster, University of Bristol (CONNECTED Network Director)
  • Prof. Susan Seal, Natural Resources Institute, University of Greenwich (CONNECTED Management Board member).

In the film Prof. Foster says: “CONNECTED is sowing the seeds of a new generation of collaborators, whose work will transform African agriculture for the next 20 to 30 years.”

You can watch below or on the CONNECTED Vimeo channel.

Waqas Khokhar talks to GARNet

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Published on: November 6, 2019

Waqas Khokhar from Canterbury Christ Church University in Kent talks to GARNet about a new publication in Frontiers in Plant Science on ‘Genome-Wide Identification of Splicing Quantitative Trait Loci (sQTLs) in Diverse Ecotypes of Arabidopsis thaliana‘. Waqas has produced a really exciting dataset that will be of great use to many other researchers!

GARNet Research Roundup: November 1st 2019

This edition of the GARNet Research Roundup begins with a pan-UK study that has identified a gene involved in starch granule formation in polyploid wheat. Second is a study from Canterbury that identifies Arabidopsis QTLs involved in alternative splicing. Third is research from Cambridge that investigates the role of the nuclear circadian oscillator on sub-cellular calcium fluctuations. The fourth paper describes the development of a computer-vision tool designed for automated measurements of wheat spikes in the field. The fifth paper is a Korean-led study that has identified a transcription factor involved in pollen development and includes co-authors from Leicester. Last is a study from the University of Warwick that has looked into light-regulated gene expression during bulb initiation in onion.


Chia T, Chirico M, King R, Ramirez-Gonzalez R, Saccomanno B, Seung D, Simmonds J, Trick M, Uauy C, Verhoeven T, Trafford K (2019) A carbohydrate-binding protein, B-GRANULE CONTENT 1, influences starch granule size distribution in a dose-dependent manner in polyploid wheat. J Exp Bot. doi: 10.1093/jxb/erz405
Open Access

Tansy Chia is lead author on this study that brings together three of the UKs major plant breeding research centres; NIAB, Rothamsted and the JIC. They take advantage of the new genomic tools and mutant populations available in wheat to characterize the complex role of the BGC1 (B-GRANULE CONTENT 1) gene during formation of B-type starch granules.


Khokhar W, Hassan MA, Reddy ASN, Chaudhary S, Jabre I, Byrne LJ, Syed NH (2019) Genome-Wide Identification of Splicing Quantitative Trait Loci (sQTLs) in Diverse Ecotypes of Arabidopsis thaliana Front Plant Sci. doi: 10.3389/fpls.2019.01160
Open Access

This work from Canterbury Christ Church University is led by Waqas Khokhar and Naeem Syed. They analysed 666 diverse Arabidopsis ecotypes to look for splicing quantitative trait loci (sQTLs)] that alter rates of alternative splicing. They identified a number of trans-sQTLs hotspots that align with known functional SNPs. This study provides the first sQTL resource across diverse ecotypes that can be used to compliment other available genome and transcriptome datasets.


Martí Ruiz MC, Jung HJ, Webb AAR (2019) Circadian gating of dark-induced increases in chloroplast- and cytosolic-free calcium in Arabidopsis. New Phytol. doi: 10.1111/nph.16280

María Carmen Martí Ruiz is lead author on this research undertaken in Alex Webb’s lab in Cambridge. They have looked at the role of the circadian clock in the control of calcium fluctuations in both cytoplasm and chloroplast stroma. They show the extent these changes are dependent on a nuclear-encoded circadian oscillator, adding a new role in sub-cellular Ca2+ signaling to the circadian machinery.


Sadeghi-Tehran P, Virlet N, Ampe EM, Reyns P, Hawkesford MJ (2019) DeepCount: In-Field Automatic Quantification of Wheat Spikes Using Simple Linear Iterative Clustering and Deep Convolutional Neural Networks. Front Plant Sci. doi: 10.3389/fpls.2019.01176
Open Access

Pouria Sadeghi-Tehran leads this theorectical study from Rothamsted Research that has developed an automated ‘DeepCount’ system for quantifying wheat spikes in the field. They use a deep convolutional neural network to test their program on field images and compare this method to other automated systems based on edge detection techniques and morphological analysis. Overall they show that this method has potential toward development of a portable and smartphone-assisted wheat-ear counting systems, that will have the associated benefits of counting accuracy and reduced labour.

https://www.frontiersin.org/articles/10.3389/fpls.2019.01176/full

Oh SA, Hoai TNT, Park HJ, Zhao M, Twell D, Honys D, Park SK (2019) MYB81, a microspore-specific GAMYB transcription factor, promotes pollen mitosis I and cell lineage formation in Arabidopsis. Plant J. doi: 10.1111/tpj.14564

Mingmin Zhao and David Twell are co-authors on this project led by Sung‐Aeong Oh and Korean colleagues. After screening pollen cell patterning mutants they have identified a role for the GAMYB transcription factor MYB81 during a narrow window prior to pollen mitosis I. They demonstrate that this protein is essential for establishing the male cell lineage in Arabidopsis pollen.


Rashid MHA, Cheng W, Thomas B (2019) Temporal and Spatial Expression of Arabidopsis Gene Homologs Control Daylength Adaptation and Bulb Formation in Onion (Allium cepa L.). Sci Rep. doi: 10.1038/s41598-019-51262-1 Open Access

This collaboration between the University of Warwick and Bangladesh Agricultural University is led by Harun Ar Rashid. They look at genetic regulation of light-dependent onion bulb initiation by growing plants under short and long days and testing the expression of known regulators of flowering time; AcFT, Ac LFY and AcGA3ox1. They also performed tissue-specific analysis to demonstrate differences in expression patterns that begin to suggest how these genes are involved in bulb initiation.

https://www.nature.com/articles/s41598-019-51262-1

Steven Spoel talks to GARNet

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Published on: October 30, 2019

Steven Spoel discusses a recent paper published in Elife entitled ‘ Dynamic ubiquitination determines transcriptional activity of the plant immune coactivator NPR1‘. This paper characterises the molecular fine-tuning that controls the transcriptional outputs and degradation of the NPR1 protein, which is a key player in the plant immune response. Thanks to Steven for the really interesting chat!

Saskia Hogenhout talks to GARNet

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Published on: October 23, 2019

Saskia Hogenhout from the John Innes Centre discusses her labs recent paper in PLOS Pathogens titled ‘Phytoplasma SAP11 effector destabilization of TCP transcription factors differentially impact development and defence of Arabidopsis versus maize‘. We also briefly discuss her international collaborations working on Phytoplasma-Crop interactions.

Fascination of Plants 2019 Success Stories

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Published on: October 21, 2019

Thank you again for all your hard work towards making this year’s Fascination of Plants Day a big success! The Success Stories 2019 report (PDF) from International Fascination of Plants Day 2019 has now been published. The FoPD coordinator Carmel Edwards thanks everyone for sending stories and photos! This will be promoted again in December when there is a full set of videos additional document. Please pass it on!

GARNet Research Roundup: October 17th 2019

This edition of the GARNet Research Roundup includes a superb selection of papers by scientists from across the UK. First is work from the Spoel lab in Edinburgh that characterizes the fine-tuning of NPR1 activity during the plant immune response. Second is work from SLCU and the University of Helsinki that is an extensive investigation into the molecular basis of cambial development. Next is research from Nottingham that looks at the importance of soil macro-structures during the growth of wheat roots.

Fourth are three papers that highlight the breadth of research occurring at the John Innes Centre. The first paper is from Enrico Coen’s lab that applies their expertise in computational modeling to leaf development in the carnivorous plant Utricularia gibba. Second is work from Saskia Hogenhout’s lab that looks at immunity to infection by Phytoplasma pathogens. Last is work from Lars Ostergaard’s lab that characterises the role of Auxin Binding promoter elements in floral development.

The seventh paper from Bristol and Glasgow looks at shade avoidance signaling via PIF5, COP1 and UVR8 whilst the eighth paper, which is from Rothamsted, demonstrates how metabolic engineering in Arabidopsis seeds can result in a high proportion of human milk fat substitute. The next paper is from the University of Durham and investigates how the composition of the Arabidopsis cell wall impacts freezing tolerance. The first author of this paper, Dr Paige Panter discusses the paper on the GARNet community podcast.

The tenth paper is from Julia Davies’s lab at the University of Cambridge and introduces an uncharacterised response to extracellular ATP signals in Arabidopsis roots. The next paper is from Mike Blatt’s group at University of Glasgow and characterises a new interaction between vesicular transport and ion channels. The penultimate entry includes co-authors from the JIC on a Chinese-led study that demonstrates improved seed vigour in wheat through overexpression of a NAC transcription factor. Finally are two methods papers taken from a special journal issue on ‘Plant Meiosis’.


Skelly MJ, Furniss JJ, Grey HL, Wong KW, Spoel SH (2019) Dynamic ubiquitination determines transcriptional activity of the plant immune coactivator NPR1. Elife. doi: 10.7554/eLife.47005
Open Access

Michael Skelly is lead author on this paper from the lab of current GARNet chair Steven Spoel. In it they investigate the mechanisms that fine-tune the function of NPR1, a key player in the plant immune response. Progressive ubiquitination of NPR1 by an E3 ligase causes both its interaction with target genes and its subsequent degradation by an E4 ligase. This latter occurrence is opposed by the deubiquitinase activity of UBP6/7, setting up a complex regulatory environment that allows the plant to rapidly response to pathogen attack.


Zhang J, Eswaran G, Alonso-Serra J, Kucukoglu M, Xiang J, Yang W, Elo A, Nieminen K, Damén T, Joung JG, Yun JY, Lee JH, Ragni L, Barbier de Reuille P, Ahnert SE, Lee JY, Mähönen AP, Helariutta Y (2019) Transcriptional regulatory framework for vascular cambium development in Arabidopsis roots. Nat Plants. doi: 10.1038/s41477-019-0522-9

This pan-European-Korean collaboration has Jing Zhang from the University of Helsinki and the Sainsbury Laboratory, University of Cambridge as first author. They use cambium cell-specific transcript profiling and follow-on network analysis to discover 62 new transcription factors involved in cambial development in Arabidopsis. This information was used to engineer plants with increased radial growth through ectopic cambial activity as well as to generate plants with no cambial activity. This understanding provides a platform for possible future improvements in production of woody biomass.


Atkinson JA, Hawkesford MJ, Whalley WR, Zhou H, Mooney SJ (2019) Soil strength influences wheat root interactions with soil macropores. Plant Cell Environ. doi: 10.1111/pce.13659
This work is led from the University of Nottingham by John Atkinson and Sacha Mooney. They use X-ray Computed Tomography to investigate a trait called trematotropism, which applies to the ability of deep rooting plants to search out macropores and avoid densely packed soil. They show root colonisation of macropores is an important adaptive trait and that strategies should be put in place to increase these structures within the natural soil environment.


Lee KJI, Bushell C, Koide Y, Fozard JA, Piao C, Yu M, Newman J, Whitewoods C, Avondo J, Kennaway R, Marée AFM, Cui M, Coen E (2019) Shaping of a three-dimensional carnivorous trap through modulation of a planar growth mechanism. PLoS Biol. doi: 10.1371/journal.pbio.3000427
Open Access

https://journals.plos.org/plosbiology/article?id=10.1371/journal.pbio.3000427

Karen Lee, Claire Bushell and Yohei Koide are co-first authors on this work led by Enrico Coen at the John Innes Centre and Minlong Cui at the Zhejiang Agriculture and Forestry University in China. This study uses 3D imaging, cellular and clonal analysis, combined with computational modelling to analyse the development of cup-shaped traps of the carnivorous plant Utricularia gibba. They identify growth ansiotrophies that result in the final leave shape that develops from an initial near-spherical form. These processes have some similarities to the polar growth seen in Arabidopsis leaves. Overall they show that ‘simple modulations of a common growth framework underlie the shaping of a diverse range of morphologies’.


Pecher P, Moro G, Canale MC, Capdevielle S, Singh A, MacLean A, Sugio A, Kuo CH, Lopes JRS, Hogenhout SA (2019) Phytoplasma SAP11 effector destabilization of TCP transcription factors differentially impact development and defence of Arabidopsis versus maize. PLoS Pathog. doi: 10.1371/journal.ppat.1008035
Open Access

This work from Saskia Hogenhout’s lab at the John Innes Centre is led by Pascal Pecher and Gabriele Moro. They look at the effect of SAP11 effectors from Phytoplasma species that infect either Arabidopsis or maize. They demonstrate that although both related versions of SAP11 destabilise plant TCP transcription factors, their modes of action have significant differences. Please look out for Saskia discussing this paper on the GARNet Community podcast next week.


Kuhn A, Runciman B, Tasker-Brown W, Østergaard L 92019) Two Auxin Response Elements Fine-Tune PINOID Expression During Gynoecium Development in Arabidopsis thaliana. Biomolecules. doi: 10.3390/biom9100526
Open Access

Andre Kuhn is first author of this research from Lars Østergaard’s lab at the John Innes Centre. They functional characterise two Auxin-responsive Elements (AuxRE) within the promotor of the PINOID gene, which are bound by the ETITIN/ARF3 Auxin Response Factor. Alteration of this AuxRE causes phenotypic changes during flower development demonstrating that even with a complex regulatory environment, small changes to cis-elements can have significant developmental consequences.


Sharma A, Sharma B, Hayes S, Kerner K, Hoecker U, Jenkins GI, Franklin KA (2019) UVR8 disrupts stabilisation of PIF5 by COP1 to inhibit plant stem elongation in sunlight. Nat Commun. doi: 10.1038/s41467-019-12369-1
Open Access

Ashutosh Sharma is first author of this UK-Spanish-Germany collaboration led by Keara Franklin at University of Bristol. They have characterised the interaction between three significant molecular players that function during the shade avoidance response in Arabidopsis; PIF5, UVR8 and COP1. In shaded conditions, UVR8 indirectly promotes rapid degradation of PIF5 through their interactions with the E3 ubiquitin ligase COP1.


van Erp H, Bryant FM, Martin-Moreno J, Michaelson LV, Bhutada G, Eastmond PJ (2019) Engineering the stereoisomeric structure of seed oil to mimic human milk fat. Proc Natl Acad Sci U S A. doi: 10.1073/pnas.1907915116

Open Access

Harrie Van Arp and Peter Eastmond at Rothamsted Research lead this extremely translational study in which they have modified the metabolic pathway for triacylglycerol (TAG) biosynthesis. By modifying the location of one biosynthesis enzyme and removing the activity of another, the fats produced in these Arabidopsis seeds are enriched to contain TAGs that are similar to those found in human milk. They propose that this technology could be used to develop a source of plant-derived human milk fat substitute.


Panter PE, Kent O, Dale M, Smith SJ, Skipsey M, Thorlby G, Cummins I, Ramsay N, Begum RA, Sanhueza D, Fry SC, Knight MR, Knight H (2019) MUR1-mediated cell-wall fucosylation is required for freezing tolerance in Arabidopsis thaliana. New Phytol. doi: 10.1111/nph.16209

Paige Panter led this work as part of her PhD at the University of Durham in the lab of Heather Knight. They characterise the role of the MUR1 protein in the control of cell wall fucosylation and how this contributes to plant freezing tolerance. Paige discusses this paper and the long history of MUR1 on the GARNet Community podcast. Please check it out!


Wang L, Stacey G, Leblanc-Fournier N, Legué V, Moulia B, Davies JM (2019) Early Extracellular ATP Signaling in Arabidopsis Root Epidermis: A Multi-Conductance Process. Front Plant Sci. doi: 10.3389/fpls.2019.01064.

Open Access

The UK-French collaboration is led by Limin Wang from Julia Davies’s lab in Cambridge. They use patch clamp electrophysiology to identify previously uncharacterized channel conductances that respond to extracellular ATP across the root elongation zone epidermal plasma membrane.


Waghmare S, Lefoulon C, Zhang B, Lileikyte E, Donald NA, Blatt MR (2019) K+ channel-SEC11 binding exchange regulates SNARE assembly for secretory traffic. Plant Physiol. doi: 10.1104/pp.19.00919

Open Access

This work from Mike Blatt’s lab in Glasgow is led by Sakharam Waghmare. They look at the interaction between SNARE proteins, which are involved in vesicular fusion and K+ channels, which help control turgor pressure during cell expansion. Through combining analysis of protein-protein interactions and electrophysiological measurement they have found that this interaction requires the activity of the regulatory protein SEC11.


Li W, He X, Chen Y, Jing Y, Shen C, Yang J, Teng W, Zhao X, Hu W, Hu M, Li H, Miller AJ, Tong Y (2019) A wheat transcription factor positively sets seed vigour by regulating the grain nitrate signal. New Phytol. doi: 10.1111/nph.16234
Wenjing Li is first author of this Chinese study that includes Yi Chen and Anthony Miller from the John Innes Centre as co-authors. This research shows that seed vigour and nitrate accumulation in wheat is regulated by the TaNAC2 transcriptions factor through its control of the TaNRT2.5 nitrate transporter. The authors suggest that both genes could be used as potential future targets to increase grain yield and nitrogen use efficiency.


The Special Issue of Methods in Molecular Biology on Plant Meiosis includes papers from the University of Cambridge, led by Christophe Lambing and the James Hutton Institute, led by Benoit Darrier.

Lambing C, Choi K, Blackwell AR, Henderson IR (2019) Chromatin Immunoprecipitation of Meiotically Expressed Proteins from Arabidopsis thaliana Flowers. Methods Mol Biol. doi: 10.1007/978-1-4939-9818-0_16
Darrier B, Arrieta M, Mittmann SU, Sourdille P, Ramsay L, Waugh R, Colas I (2019) Following the Formation of Synaptonemal Complex Formation in Wheat and Barley by High-Resolution Microscopy. Methods Mol Biol. doi: 10.1007/978-1-4939-9818-0_15

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