organism: Homo sapiens
description: Protocols: Cells were maintained in 60mm dishes at 37 °C in a humidified atmosphere at 5% CO2 in DMEM 4.5g/L Glucose with UltraGlutamine media supplemented with 10% of Tet-free FBS (Clontech) and 1% penicillin/streptomycin. Cells were washed twice with PBS and added 700µL of Trizol and continue with the extraction protocol. Total RNA from HEK293 and HeLa was extracted using the miRNeasy Mini kit. Total RNA was resolved on 15% Novex TBE-urea gels and size-selected for 60-100 nt fragments. The recovered material was then alkaline hydrolyzed (10 mM sodium carbonate and 10 mM sodium bicarbonate) for 10 min at 60°C. Treated following the protocol of hydro-tRNAseq (Gogakos et al. 2017) The resulting RNA was de-phosphorylated with Antarctic Phosphatase (New England Biolabs) at 37°C for 1 h. De-phosphorylated RNA was purified with an RNeasy MinElute spin column and re-phosphorylated with polynucleotide kinase (NEB). PNK-treated tRNAs were purified with an RNeasy MinElute spin column, adaptor-ligated, reverse-transcribed, and PCR-amplified for 14 cycles. The resulting cDNA was purified using a QIAQuick PCR Purification Kit.
attributes.tag: ENA first public
attributes.value: 2021-03-23
attributes.tag: ENA last update
attributes.value: 2020-12-11
attributes.tag: External Id
attributes.value: SAMEA7705637
attributes.tag: INSDC center alias
attributes.value: CENTER FOR GENOMIC REGULATION (CRG)
attributes.tag: INSDC center name
attributes.value: CENTER FOR GENOMIC REGULATION (CRG)
attributes.tag: INSDC first public
attributes.value: 2021-03-23T08:57:08Z
attributes.tag: INSDC last update
attributes.value: 2020-12-11T15:40:18Z
attributes.tag: INSDC status
attributes.value: public
attributes.tag: Submitter Id
attributes.value: E-MTAB-9905:CAT1
attributes.tag: age
attributes.value: 62
attributes.tag: broker name
attributes.value: ArrayExpress
attributes.tag: cell line
attributes.value: HaCaT
attributes.tag: cell type
attributes.value: keratinocyte
attributes.tag: common name
attributes.value: human
attributes.tag: developmental stage
attributes.value: adult
attributes.tag: disease
attributes.value: normal
attributes.tag: genotype
attributes.value: wild type genotype
attributes.tag: organism part
attributes.value: skin epidermis
attributes.tag: sample name
attributes.value: E-MTAB-9905:CAT1
attributes.tag: sex
attributes.value: male