|
| Status |
Public on May 01, 2015 |
| Title |
GRA23-knockout |
| Sample type |
SRA |
| |
|
| Source name |
infected BMMs
|
| Organisms |
Mus musculus; Toxoplasma gondii RH |
| Characteristics |
pathogen: Toxoplasma gondii strain Rh
mouse strain: C57BL/6
tissue: Bone marrow
cell type: macrophages
age: week 4-6 mice
toxoplasma gondii genotype: dKu80/dGRA23/+HXGPRT
|
| Treatment protocol |
Cells were infected with indicated Toxplasma strains for 4 hours at an intended MOI of 5
|
| Growth protocol |
Cells were grown in R20 media (RPMI Glutamax +20% L929 sup+ 10% FCS +10uM HEPES +50uM B-ME + Pen.Strep )
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was isolated using QIAGEN® RNeasy plus kit according the manufacturer’s instructions. Sequencing libraries were prepared according to Illumina RNA-Seq library kit with minor modifications. Briefly, mRNA was isolated using Dynabeads® mRNA Purification Kit (Invitrogen) followed by fragmentation (Ambion) and ethanol precipitation. First and second strand synthesis were performed followed by end repair, A-tailing, paired end adapter ligation and size selection on agarose gels. 200-400 bp dsDNA was enriched by 15 cycles of PCR with Phusion® High-Fidelity DNA Polymerase (NEB) followed by gel purification of ~250 bp fragments from the amplified material. Amplified libraries were sequenced on an Illumina GAII or GAIIx sequencer, alternatively.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2000 |
| |
|
| Description |
as decribed in http://www.ncbi.nlm.nih.gov/pubmed/23231500
|
| Data processing |
Illumina Offline BaseCaller1.9.3 software used for basecalling.
Sequence reads were mapped to either mm9 whole genome using Tophat 2.0.4 with the following parameters --genome-read-mismatches 2 --read-mismatches 3 --min-intron-length 70 --max-intron-length 500000 --solexa1.3-quals --no-coverage-search --segment-length 20 --mate-inner-dist 220 --mate-std-dev 30 --min-anchor-length 4 --b2-sensitive or to ME49v8.0 whole genome using Tophat 2.0.4 --no-coverage-search --max-coverage-intron 2000 --max-segment-intron 2000 --segment-length 20 --mate-std-dev 80 -a 8 -i 70 -I 2000
Gene expression was calculated as Fragments per Kilobase of modeled exon per Million mapped reads (FPKM) using Cufflinks 2.0.2
Genome_build: mm9 or ME49v8.0
Supplementary_files_format_and_content: A tab-delimited text file that includes FPKM values for each Sample.
|
| |
|
| Submission date |
Feb 17, 2015 |
| Last update date |
May 15, 2019 |
| Contact name |
Daniel A Gold |
| Organization name |
MIT
|
| Department |
Biology
|
| Lab |
Saeij Lab
|
| Street address |
77 Massachusetts Ave 68-264d
|
| City |
Cambridge |
| State/province |
Massachusetts |
| ZIP/Postal code |
02139 |
| Country |
USA |
| |
|
| Platform ID |
GPL19788 |
| Series (1) |
| GSE65980 |
Toxoplasma proteins GRA17 and GRA23 mediate the movement of small molecules between the host and the parasitophorous vacuole. |
|
| Relations |
| BioSample |
SAMN03351438 |
| SRA |
SRX878813 |
