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| Status |
Public on Jul 10, 2012 |
| Title |
shApoB (small RNA) |
| Sample type |
SRA |
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| Source name |
shApoB
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| Organism |
Homo sapiens |
| Characteristics |
cell type: HEK293T
cell source: ATCC CRL-11268™
tissue: Kidney
genotype: wild type
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| Extracted molecule |
total RNA |
| Extraction protocol |
HEK293T cells were transfected with 4 µg shApoB- or miApoB-expression plasmids using Lipofectamine 2000 reagent using manufacturers protocol and total RNA was isolated from cells 48 hr post-transfection using Trizol. Small RNA sequencing libraries were generated using high-quality total RNA as input and the NEXTflex Small RNA Sequencing kit (Bioo Scientific, USA). Briefly, the small RNA species were subjected to ligation with 3' and 5' RNA adapters, first strand reverse transcription, and PCR amplification. Sample-specific barcodes were introduced in the PCR step. The PCR products were separated on TBE-PAGE electrophoresis and the expected band around 140-160bp was recovered for each sample. The libraries were multiplexed, clustered, and sequenced on an Illumina HiSeq 2000.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
size fractionation |
| Instrument model |
Illumina HiSeq 2000 |
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| Data processing |
Illumina Casava1.8.2 software used for basecalling.
The raw sequencing data produced was processed removing the sequence reads which were of too low quality (only "passing filter" reads were selected), and discarding reads that aligned against the PhiX control. In total, we generated 16016701 reads for sample miApoB, and 15843150 reads for sample shApoB.
The obtained reads were adaptor-trimmed, which decreased the average read size from ~36bp to ~25bp. The custom adapter sequenced used for trimming all the bases extending 5’ was: GTGACTGGAGTTCC-TTGGCACCCGAGAATTCCA.
Next, both data sets from shApoB and miApoB samples were grouped based on the match to the reference sequence and the obtained unique small RNAs were aligned to the sequence of pre-miApoB: GATCCTGGAGGCTTGC-TGAAGGCTGTATGCTGATGGACAGGTCAATCAATCTTGTTTTGGCCACTGACTGACAAGATTGAGACCTGTCCATCAGGACACAAGGCCTGTTACTAGCACTCACATGGAACAAATGGCCCAGATCTGGCCGCAG or shApoB: GATCCCCGATTGATTGACCTGTCCATTTCAAGAGAATGGACAGGTCAATCAATC-TTTTTCAGCTT sequence, respectively.
Supplementary_files_format_and_content: Spreadsheet including reads abundancy. Figure 1 and showing alliingment with references miApoB and shApoB sequences
Genome_build: miRBase Release 18: November 2011
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| Submission date |
Jul 09, 2012 |
| Last update date |
May 15, 2019 |
| Contact name |
Piotr Maczuga |
| E-mail(s) |
p.maczuga@uniqure.com
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| Organization name |
UniQure
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| Street address |
Meibergdreef 61
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| City |
Amsterdam |
| State/province |
Nord Holland |
| ZIP/Postal code |
1105BA |
| Country |
Netherlands |
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| Platform ID |
GPL11154 |
| Series (1) |
| GSE39187 |
Embedding siRNA sequences targeting Apolipoprotein B100 in shRNA and miRNA scaffolds results in differential processing and in vivo efficacy |
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| Relations |
| SRA |
SRX158351 |
| BioSample |
SAMN01086231 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM957705_shApoB.txt.gz |
201.9 Kb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
| Processed data provided as supplementary file |
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