Virus-Induced Gene Silencing: Methods and Protocols by Matthias Lange, Aravinda L. Yellina, Svetlana Orashakova
By Matthias Lange, Aravinda L. Yellina, Svetlana Orashakova (auth.), Annette Becker (eds.)
Plants are extraordinary organisms to review, a few are very important resources for prescription drugs, and others may help to explain molecular mechanisms required for a plant’s improvement and its interactions with the biotic or abiotic atmosphere. sensible genomics is significantly lagging at the back of the rate of genome sequencing as high-throughput gene functionality assays are tricky to layout, in particular for non-model crops. Bioinformatics instruments are beneficial for gene id and annotation yet are of restricted worth for predictions referring to gene features as gene capabilities are exposed top via experimental methods. Virus-Induced-Gene-Silencing (VIGS) is a simple to exploit, quickly, and trustworthy technique to in achieving down rules of goal gene expression. Virus-Induced Gene Silencing: tools and Protocols provides distinctive protocols for VIGS experiments in different plant species together with version and non-model crops. additionally incorporated during this booklet are lately constructed protocols for VIGS-derived microRNA creation within the plant or protein over expression, in addition to chapters dedicated to summarizing the molecular mechanisms of VIGS motion and the vector structures constructed up to now. Written within the winning Methods in Molecular Biology™ sequence layout, chapters contain introductions to their respective themes, lists of the mandatory fabrics and reagents, step by step, without difficulty reproducible protocols, and notes on troubleshooting and heading off recognized pitfalls.
Authoritative and simply available, Virus-Induced Gene Silencing: equipment and Protocolsservesas a priceless source for researchers from diversified fields of plant biology attracted to experimental methods to interpreting gene functions.
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Extra resources for Virus-Induced Gene Silencing: Methods and Protocols
Ding et al. (21) cloned and modified the fescue strain of BMV (F-BMV, isolated from Festuca arundinacea) to function as a VIGS vector. A clone of genomic RNA 3 representing the Russian strain of BMV (R-BMV) was used with genomic clones representing RNAs 1 and 18 H. Ramanna et al. 2 of F-BMV to allow easy foreign gene insertion (due to a unique HindIII restriction site in the R-BMV RNA 3 clone) and infectivity to rice (due to the use of clones representing F-BMV RNAs 1 and 2 (21)). Foreign gene fragments were inserted downstream of the coat protein stop codon within the cDNA representing RNA 3.
It is also worthwhile to identify new viral vectors that are not transmitted by insect vectors and seed so that studies can be conducted under less stringent governmental regulations. S. S. Nelson, unpublished). PMV is a single-stranded RNA virus (genus Panicovirus) and causes mild mosaic symptoms in many monocotyledonous species including S. italica and maize cultivar Oh28 (86, 87). PMV is readily transmitted among plants through mechanical inoculation but not through insect vectors and seed (86, 88).
Tai YS, Bragg J, Edwards MC (2005) Virus vector for gene silencing in wheat. Biotechniques 39:310–312 20. Meng Y, Moscou MJ, Wise RP (2009) Blufensin1 negatively impacts basal defense in response to barley powdery mildew. Plant Physiol 149:271–285 21. Ding XS, Schneider WL, Chaluvadi SR et al (2006) Characterization of a Brome mosaic virus strain and its use as a vector for gene silencing in monocotyledonous hosts. Mol Plant Microbe Interact 19:1229–1239 22. Pacak A, Strozycki PM, Barciszewska-Pacak M et al (2010) The Brome mosaic virus-based recombination vector triggers a limited gene silencing response depending on the orientation of the inserted sequence.