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General Information
Name
mE_mRNA_A_4d_Caffeine_2.5mg/ml
Species
D. melanogaster
Result type
FlyBase ID
FBlc0000244
Project
Data Provider
Title
RNA-Seq coverage profile of D. melanogaster, Oregon-R-modENCODE strain, adult (4 day), 2.5mg/ml caffeine treatment.
Status
Current
Accessions
    Biosample Source
    Overview
    Sex
    Tissue isolated
    Other tissues studied
    Cell component
    Cell line
    Key genes
    Sample preparation

    Newly eclosed adults were aged for 2 days, then exposed to caffeine (2.5mg/ml in 10% sucrose solution) for 48 hours; 41.9% lethality was observed. Survivors were flash frozen in liquid nitrogen and stored at -80oC prior to RNA preparations.

    Biosamples analyzed by this result (1)
    Biosample
    Type
    Title
    D. melanogaster, Oregon-R-modENCODE strain, adult (4 day), 2.5mg/ml caffeine treatment, source for RNA.
    Data Analyzed
    Key genes
    Protocol

    Total RNA was isolated using Trizol, then Qiagen RNeasy spin columns. mRNA was isolated using Dynal oligo(dT) beads, then fragmented using divalent cations under elevated temperature, followed by sequential ligation of RNA linkers to the 5’ and 3’ ends. Next, reverse transcription was performed using a primer complementary to the 3’ linker and PCR was performed using primers complementary to both linkers. ~ 300 bp fragments were isolated from an agarose gel and gel-purified again.

    Mode of Assay

    Read length (bases):100

    The samples were quantitated using a Nanodrop, and loaded onto a flow cell for cluster generation and sequenced on an Illumina Genome Analyzer II using either single read or paired end protocols (Illumina).

    Raw Data Analyzed (1)
    Assay / Reagent collection
    Type
    Title
    RNA-Seq of D. melanogaster, Oregon-R-modENCODE strain, adult (4 day), 2.5mg/ml caffeine treatment.
    Processed Data Analyzed (0)
    Result
    Type
    Title
    Analysis
    Reference Genome
    Reference Annotation
    Data analysis

    Reads were aligned to Dmel_Release_6 using the STAR aligner v2.3.0e (Linux x86_64) with default parameters on the FASTQ files to generate multiply-mapped BAM files. These were filtered to include reads with only 1 aligned hit ( NH:i:1 attribute) to generate uniquely-mapped BAM files. A custom script was used to convert BAM files into bedgraph files (bam2bedgraph.cc).

    Comments
    Associated Data
    Size
    Number of unique reads.
    Files
    Additional Information

    The RNA-seq profiles displayed by FlyBase in GBrowse and used for RPKM calculation can be accessed at the FTP link below as .wig files. Please take note of how these FlyBase .wig files represent data for a contiguous sequence of bases with the same signal value. The value is declared only for the first position of that region, and applies to all positions that follow (these are not explicitly listed) until a new value at a new base position is declared.

    Synonyms and Secondary IDs (6)
    Reported As
    Symbol Synonym
    Caffeine Treatment_2.5 mg/ml_Four Day Adults
    Caffeine_2.5mg_AdMixed_4d
    Cherbas_750 Cherbas_751
    mE_mRNA_A_4d_Caffeine_2.5mg/ml
    Name Synonyms
    RNA-Seq coverage profile of D. melanogaster, Oregon-R-modENCODE strain, adult (4 day), 2.5mg/ml caffeine treatment.
    Secondary FlyBase IDs
      References (5)