Tuesday, November 1, 2016

Methods reviews for in vivo RNAi screening in Drosophila

Four! RNAi-relevant methods papers in the recent Drosophila-focused issue of Methods in Molecular Biology:

Billmann M, Boutros M. Methods for High-Throughput RNAi Screening in Drosophila Cells. Methods Mol Biol. 2016;1478:95-116. PMID: 27730577.

Kaya-Çopur A, Schnorrer F. A Guide to Genome-Wide In Vivo RNAi Applications in Drosophila. Methods Mol Biol. 2016;1478:117-143. PMID: 27730578.

Zhou J, Tong C. Design and Methods of Large-Scale RNA Interference Screens in Drosophila. Methods Mol Biol. 2016;1470:163-9. PMID: 27581292. Chen X, Xu L.

Genome-Wide RNAi Screening to Dissect the TGF-β Signal Transduction Pathway. Methods Mol Biol. 2016;1344:365-77. PMID: 26520138.

Monday, October 31, 2016

RNAi used to induce ROS stress in a study of oogenesis

Perkins AT, Das TM, Panzera LC, Bickel SE. Oxidative stress in oocytes during midprophase induces premature loss of cohesion and chromosome segregation errors. Proc Natl Acad Sci U S A. 2016 Oct 17. pii: 201612047. PMID: 27791141.

From the abstract: "... A growing body of evidence suggests that meiotic cohesion deteriorates as oocytes age ... One hallmark of aging cells is an increase in oxidative damage caused by reactive oxygen species (ROS). Therefore, increased oxidative damage in older oocytes may be one of the factors that leads to premature loss of cohesion and segregation errors. To test this hypothesis, we used an RNAi strategy to induce oxidative stress in Drosophila oocytes and measured the fidelity of chromosome segregation during meiosis. ..."

Tick tock! Screening the clock.

Agrawal P, Hardin PE. An RNAi Screen to Identify Protein Phosphatases that Function Within the Drosophila Circadian Clock. G3 (Bethesda). 2016 Oct 26. PMID: 27784754.

From the abstract: "Circadian clocks in eukaryotes keep time via cell-autonomous transcriptional feedback loops. ... To identify phosphatases that play a role in the clock, we screened clock cell-specific RNA interference (RNAi) knockdowns of all annotated protein phosphatases and protein phosphatase regulators in Drosophila for altered activity rhythms. ... Additional RNAi lines, transposon inserts, overexpression and loss-of-function mutants were tested to independently confirm these RNAi phenotypes. ... 15 viable protein phosphatases remain for future studies. These candidates are expected to reveal novel features of the circadian timekeeping mechanism in Drosophila that are likely to be conserved in all animals including humans."

Thursday, October 13, 2016

Methods publications relevant to cell and in vivo RNAi

Methods in Molecular Biology has recently published papers relevant to Drosophila cell culture, cell-based RNAi, and in vivo RNAi.

Debec A, Megraw TL, Guichet A. Methods to Establish Drosophila Cell Lines. Methods Mol Biol. 2016;1478:333-351. PubMed PMID: 27730593

Billmann M, Boutros M. Methods for High-Throughput RNAi Screening in Drosophila Cells. Methods Mol Biol. 2016;1478:95-116. PubMed PMID: 27730577.

Kaya-Çopur A, Schnorrer F. A Guide to Genome-Wide In Vivo RNAi Applications in Drosophila. Methods Mol Biol. 2016;1478:117-143. PubMed PMID: 27730578.

Monday, September 12, 2016

Drosophila CRISPR protocols series

Thinking fly CRISPR? Series of methods papers out in CSH Protocols.

Housden BE, Perrimon N. Cas9-Mediated Genome Engineering in Drosophila melanogaster. Cold Spring Harb Protoc. 2016 Sep 1;2016(9):pdb.top086843. PMID: 27587786.

Housden BE, Perrimon N. Detection of Indel Mutations in Drosophila by High-Resolution Melt Analysis (HRMA). Cold Spring Harb Protoc. 2016 Sep 1;2016(9):pdb.prot090795. PMID: 27587781.

Housden BE, Perrimon N. Design and Generation of Donor Constructs for Genome Engineering in Drosophila. Cold Spring Harb Protoc. 2016 Sep 1;2016(9):pdb.prot090787. PMID: 27587780.

Housden BE, Hu Y, Perrimon N. Design and Generation of Drosophila Single Guide RNA Expression Constructs. Cold Spring Harb Protoc. 2016 Sep 1;2016(9):pdb.prot090779. PMID: 27587779.

Tuesday, September 6, 2016

Housden BE, Hu Y, Perrimon N. Design and Generation of Drosophila Single Guide RNA Expression Constructs. Cold Spring Harb Protoc. 2016 Sep 1;2016(9):pdb.prot090779. PMID: 27587779.

From the abstract: "... Here, we provide a step-by-step protocol for designing sgRNA target sites using the Drosophila RNAi Screening Center (DRSC) Find CRISPRs tool (version 2). We also describe the generation of sgRNA expression plasmids for the use in cultured Drosophila cells or in vivo. Finally, we discuss specific design requirements for various genome engineering applications."

Methods paper -- in vivo RNAi screens

Zhou J, Tong C. Design and Methods of Large-Scale RNA Interference Screens in Drosophila. Methods Mol Biol. 2016;1470:163-9. PMID: 27581292.

From the abstract: "... Here, we discuss methods for the design and performance of a large-scale in vivo RNAi screen in Drosophila. Furthermore, methods for the validation of results and analysis of data will be introduced."