title: 919R1
organism: Hordeum vulgare subsp. vulgare
description: Protocols: Barley plants of Hordeum vulgare cv. Golden Promise were used. Grains were germinated in soil and grown at 22ºC under a 16 h light/8 h darkness photoperiod for 7 days in Sanyo MLR-350-H chambers. Barley transgenic lines overexpressing or silencing the HvPap-1 gene (OE Pap1 and KD Pap1, respectively) were generated using Agrobacterium-mediated gene transfer on immature embryos and homozygous material obtained by double haploid technology. Transgenic barley plants ubiquitously over-expressing the HvPap-1 gene were generated using p6U and p6d35s binary vectors. Silencing of the HvPap-1 gene was performed using the artificial microRNA (amiRNA) technology. Total RNA was extracted from frozen barley leaves by the phenol/chloroform method, followed by precipitation with 8 M LiCl and digested with DNase (Promega). Using poly-T oligo-attached magnetic beads, mRNAs were purified from the total RNA. Then, the mRNAs were fragmented and cDNA was synthesized using random hexamer-primers, DNA polymerase I and RNase H. The double-stranded cDNAs were purified with magnetic beads and ligated to adaptors for Illumina sequencing. The quality and quantity of the library was verified using an Agilent 2100 Bioanalyzer and an ABI StepOnePlus Real-Time PCR system, respectively.
attributes.tag: ENA first public
attributes.value: 2017-09-29
attributes.tag: ENA last update
attributes.value: 2017-08-10
attributes.tag: External Id
attributes.value: SAMEA104209661
attributes.tag: INSDC center alias
attributes.value: Centro de Biotecnologia y Genomica de Plantas, Universidad Politecnica de Madrid (UPM) - Instituto Nacional de Investigacion y Tecnologia Agraria y Alimentaria (INIA)
attributes.tag: INSDC center name
attributes.value: Centro de Biotecnologia y Genomica de Plantas, Universidad Politecnica de Madrid (UPM) - Instituto Nacional de Investigacion y Tecnologia Agraria y Alimentaria (INIA)
attributes.tag: INSDC first public
attributes.value: 2017-09-29T17:01:53Z
attributes.tag: INSDC last update
attributes.value: 2017-08-10T14:12:56Z
attributes.tag: INSDC status
attributes.value: public
attributes.tag: Submitter Id
attributes.value: E-MTAB-5963:919R1
attributes.tag: age
attributes.value: 7
attributes.tag: broker name
attributes.value: ArrayExpress
attributes.tag: common name
attributes.value: domesticated barley
attributes.tag: cultivar
attributes.value: Golden Promise
attributes.tag: genotype
attributes.value: Pap1 overexpression
attributes.tag: organism part
attributes.value: leaf
attributes.tag: sample name
attributes.value: E-MTAB-5963:919R1
attributes.tag: sub_species
attributes.value: vulgare