 |
 |
GEO help: Mouse over screen elements for information. |
|
| Status |
Public on Apr 03, 2012 |
| Title |
Wholeflies_ED777_Female_DNASeq_R1 |
| Sample type |
SRA |
| |
|
| Source name |
Wholeflies, 5 day mated adults
|
| Organism |
Drosophila melanogaster |
| Characteristics |
gender: Female
strain: Df(2L)ED777/+
tissue: whole flies
development stage: 5 day old mated adults
|
| Treatment protocol |
We outcrossed the balancer chromosome using females of w1118 and collected 30 -100 adult, 5 day old sexed flies containing Df/+ for DNA collection.
|
| Growth protocol |
Flies were grown under constant light, temperature, and humidity (25°C; 60% relative humidity) on San Diego Stock Center cornmeal media (DSSC, 2008)
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Flies were homogenized in 200 μl Buffer A (100mM Tris-HCl, pH 7.5, 100 mM EDTA, 100 mM NaCl, 0.5% SDS), then an additional 200 μl of Buffer A was added followed by homogenization. Samples were incubated at 65°C for 30 minutes, then 800 μl LiCl/KAc solution (1 part 5 M KAc: 2.5 parts 6 M LiCl) was added. Samples were spun for 15 minutes at room temperature at 14000 rpm in a microcentrifuge. One ml of clear supernatant was removed from samples and placed in a new 1.5 ml Eppendorf tube, then 600 μl of isopropanol was added, and the samples were spun for 15 minutes at room temperature at 14000 rpms. Supernatant was removed and the DNA pellet was washed with equal volume 70% EtOH. Pellets were dried for 5-10 minutes, then re-suspended in 150 μl TE Buffer. Five μg genomic DNA in 200 μl TE buffer was sonicated on high for 30 sec on and 30 sec off for a total of 30 min using a Bioruptor UCD-200 (Diagenode, Sparta New Jersey). The fragmented DNA was then purified using a QIAGEN QIAquick PCR purification kit. We prepared 1-1.5 μg of fragmented DNA and followed the Illumina protocols for paired-end sequencing library preparation or following standard protocols from Illumina TruSeq kits. For the samples we sequenced with TruSeq, we performed multiplexing of samples for sequencing on a Illumina HiSeq 2000 instrument with four samples per lane in six lanes and six samples per lane in two lanes of a flowcell.
|
| |
|
| Library strategy |
OTHER |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
Illumina HiSeq 2000 |
| |
|
| Data processing |
Libraries were sequenced on an Illumina GA II or multiplexed on an Illumina HiSeq 2000 to generate 36 bp single end reads and those reads passing the Illumina quality filter were mapped with Bowtie 0.12.7 using -v 2 -m 1 to assembly 5 of the Drosophila melanogaster genome excluding chromosome Uextra (dm3 from UCSC Genome Broswer). We used the unique mapping reads to generate coverage plots using the genomeCoverageBed in BEDTools. Normalized coverage, along with the start and stop positions of Df/+ regions were visualized on the UCSC Genome Browser. Deficient regions were characterized by a loss of read coverage in the region and most were unambiguously definable. Results for 20 Df(2L)EDs were as expected (e.g. nucleotide level breakpoint accuracy and approximate 2-fold differences in DNA-Seq read density, but we did observe an additional deficiency region in the Df(2L)ED478 line, which we named Df(2L)Hsp60c.
|
| |
|
| Submission date |
Aug 19, 2011 |
| Last update date |
May 15, 2019 |
| Contact name |
Brian Oliver |
| E-mail(s) |
[email protected]
|
| Phone |
301-204-9463
|
| Organization name |
NIDDK, NIH
|
| Department |
LBG
|
| Lab |
Developmental Genomics
|
| Street address |
50 South Drive
|
| City |
Bethesda |
| State/province |
MD |
| ZIP/Postal code |
20892 |
| Country |
USA |
| |
|
| Platform ID |
GPL13304 |
| Series (2) |
| GSE31407 |
Drosophila DrosDel deficiency and diploid control flies |
| GSE31550 |
DNA-Seq of DrosDel deficiency lines and w1118 lines |
|
| Relations |
| SRA |
SRX092547 |
| BioSample |
SAMN00710492 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM783123_ED777-F_bowtiev2m1.bam |
634.0 Mb |
(ftp)(http) |
BAM |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
|
|
|
|
 |
