organism: Homo sapiens
description: Protocols: RNA purity, integrity and concentration were determined using an Agilent 2100 Bioanalyzer (Agilent Technologies, Inc, USA). Only RNA with a RIN value of 8.0 or higher and a ratio (28s/18s) of above 1.8 was taken forward for sample preparation. Samples were processed for library constructions following the manufactory instructions (Illumina Tru Seq Stranded Total RNA sample preparation v2 Guide, Part #15031048 Rev.E October 2013 - “Low sample protocol”)*. In brief, 0.5 µg of total RNA from each tissue were depleted for cytoplasmic rRNA using the Ribo-Zero ribosomal reduction chemistry, chemically fragmented for 8 min at 94oC, and processed for 1st strand synthesis cDNA and then 2nd strand synthesis using dUTP instead of dTTP. After some purification steps, the cDNA was then end-repaired, purified, adenylated at the 3'-ends, and purified before adding the indexed adaptor sequences using the TruSeq Stranded LT Kit Index set A. Each library preparation was then enriched by 10 cycles of PCR, purified and finally validated in regards to size and concentration. For sizing, libraries were analyzed on the Agilent 2100 Bioanalyzer using a DNA 1000 kit from Agilent Technologies. The Libraries were quantified by qPCR using the KaPa Library quantification Kits (KaPa Biosystems, Cat KK4824). Samples were pooled in sets of 12 libraries, and a final concentration of 16 pM denatured library was used for paired-end deep sequencing using an Illumina HiSeq1500.
attributes.tag: ENA first public
attributes.value: 2016-08-25
attributes.tag: ENA last update
attributes.value: 2016-07-25
attributes.tag: External Id
attributes.value: SAMEA4355087
attributes.tag: INSDC center alias
attributes.value: University of Southern Denmark Department of Biochemistry and Molecular Biology
attributes.tag: INSDC center name
attributes.value: University of Southern Denmark Department of Biochemistry and Molecular Biology
attributes.tag: INSDC first public
attributes.value: 2016-08-25T17:01:19Z
attributes.tag: INSDC last update
attributes.value: 2016-07-25T12:22:57Z
attributes.tag: INSDC status
attributes.value: public
attributes.tag: Submitter Id
attributes.value: E-MTAB-36641469444294:HeLa_SMN_KD_1
attributes.tag: broker name
attributes.value: ArrayExpress
attributes.tag: common name
attributes.value: human
attributes.tag: genotype
attributes.value: adenocarcinoma
attributes.tag: organism part
attributes.value: HeLa cell culture
attributes.tag: sample name
attributes.value: E-MTAB-36641469444294:HeLa_SMN_KD_1
attributes.tag: sex
attributes.value: female
attributes.tag: treatment
attributes.value: SMN siRNA