Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

A genome-scale shRNA resource for transgenic RNAi in Drosophila

ABSTRACT: Existing transgenic RNAi resources in Drosophila utilize the expression of long double stranded hairpin RNAs and are powerful tools for the identification and characterization of genes involved in various processes. However, they are ineffective to knockdown genes during oogenesis, an important model system for the study of numerous fundamental questions in biology. Here, we show that short hairpin RNAs (shRNAs), modeled on an endogenous microRNA, are extremely effective at silencing gene expression during oogenesis. We also describe our progress towards building a genome-wide shRNA resource. Total RNA was isolated using Trizol reagent (Invitrogen) and size-fractionated by PAGE. These were independently processed and sequenced using the Illumina GAII platform. In total, three libraries were analyzed.

ORGANISM(S): Drosophila melanogaster

SUBMITTER: Benjamin Czech

PROVIDER: E-GEOD-27039 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

A genome-scale shRNA resource for transgenic RNAi in Drosophila.

Ni Jian-Quan JQ Zhou Rui R Czech Benjamin B Liu Lu-Ping LP Holderbaum Laura L Yang-Zhou Donghui D Shim Hye-Seok HS Tao Rong R Handler Dominik D Karpowicz Phillip P Binari Richard R Booker Matthew M Brennecke Julius J Perkins Lizabeth A LA Hannon Gregory J GJ Perrimon Norbert N

Nature methods 20110403 5

Existing transgenic RNAi resources in Drosophila melanogaster based on long double-stranded hairpin RNAs are powerful tools for functional studies, but they are ineffective in gene knockdown during oogenesis, an important model system for the study of many biological questions. We show that shRNAs, modeled on an endogenous microRNA, are extremely effective at silencing gene expression during oogenesis. We also describe our progress toward building a genome-wide shRNA resource. ...[more]

PMID: 21460824

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Project description:Wheat streak mosaic virus (WSMV) and Triticum mosaic virus (TriMV) are type members of Tritimovirus and Poacevirus genera, respectively, in the family Potyviridae, and are transmitted by wheat curl mites. Co-infection of these two viruses causes synergistic interaction with increased virus accumulation and disease severity in wheat. In this study, we examined the effects of synergistic interaction between WSMV and TriMV on endogenous small (s) RNAs and virus-specific small interfering RNAs (vsiRNAs) in susceptible (Arapahoe) and temperature-sensitive resistant (Mace) wheat cultivars at 27C and 18C. Single- and double-infections in wheat caused a shift in the profile of endogenous sRNAs from 24 nt being the most predominant in healthy plants to 21 nt in infected wheat. Additionally, we report high-resolution vsiRNA maps of WSMV and TriMV in singly- and doubly-infected wheat cultivars Arapahoe and Mace at 18C and 27C. Massive amounts of 21 and 22 nt vsiRNA reads were accumulated in Arapahoe at both temperatures and in Mace at 27C but not at 18C. The plus- and minus-sense vsiRNAs were distributed throughout the genomic RNAs in Arapahoe at both temperature regimens and in Mace at 27C, although some regions of genomic RNAs serve as hot-spots with an excessive number of vsiRNAs. The positions of vsiRNA peaks were conserved among wheat cultivars Arapahoe and Mace, suggesting that Dicer-like enzymes of susceptible and resistant wheat cultivars are similarly accessed the genomic RNAs of WSMV and TriMV. Additionally, several cold-spot regions were found in the genomes of TriMV and WSMV with no or a few vsiRNAs, indicating that certain regions of WSMV and TriMV genomes are not accessible to Dicer-like enzymes. The high-resolution map of endogenous and vsiRNAs from wheat cultivars synergistically infected with WSMV and TriMV at two temperature regimens form a foundation for understanding the virus-host interactions, effect of synergistic interactions on host defense mechanisms, and virus resistance mechanisms in wheat. Small RNA was sequenced from two wheat cultivars (Mace and Araphahoe), at two temperatures 18C and 27C, for healthy (control/uninfected), infected with wheat streak mosaic virus (WSMV), infected with Triticum mosaic virus (TriMV), and a double-infecttion of WSMV and TriMV.

Dataset Information

A genome-scale shRNA resource for transgenic RNAi in Drosophila

Publications

A genome-scale shRNA resource for transgenic RNAi in Drosophila.

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