organism: Zea mays
description: Protocols: GA2OX::PLA1 transgenic plants and their none-transgenic siblings were grown in the field in Wetteren and in the growth chamber. In the field, we sampled three plants per plot (three plots were sown per genotype), resulting in three pools each consisting of three plants. Every pool represented a different subplot. In the growth chamber, the plants were sampled randomly across the population, with three pools, each consisting of three plants. For each plant, the day of leaf 4 appearance was marked and the samples were taken two (and six) days after leaf 4 appeared. The seedlings were cut just above soil level and the growing fourth leaf was dissected out and the younger leaves inside leaf 4 were removed. Using a ruler we sampled the basal cm of leaf 4 (division zone) for RNA preparation. The samples were snap frozen and transported to the lab for storage at -80 degrees and further processing. The dissection of leaf 4 is shown in the following movie http://www.psb.ugent.be/~hinel/maisfilmpje.mp4. The basal cm of leaf 4 was sampled. The plants were grown in the field in May-June 2016 and in the growth chamber (relative humidity (55%), temperature (24 °C day/18 °C night) and 16 h/8 h (day/night) cycle). Total RNA was extracted using the guanidinium thiocyanate-phenol-chloroform extraction method using TRI-reagent (Thermo Fisher Scientific) followed by DNA digestion using the RQ1 RNase-free DNase kit (Promega). Total RNA was sent to GATC Biotech for RNA sequencing. Library preparation was done using the NEBNext Kit (Illumina). In brief, purified poly(A)-containing mRNA molecules were fragmented, randomly primed strand-specific cDNA was generated and adaptors were ligated. After quality control using an Advanced Analytical Technologies Fragment Analyzer, clusters were generated through amplification using cBOT (Cluster Kit v4, Illumina).
attributes.tag: ENA-FIRST-PUBLIC
attributes.value: 2019-10-16T17:04:25Z
attributes.tag: ENA-LAST-UPDATE
attributes.value: 2019-06-27T12:41:31Z
attributes.tag: External Id
attributes.value: SAMEA5740776
attributes.tag: INSDC center name
attributes.value: Department of Plant Biotechnology and Bioinformatics, Ghent University, Gent, Belgium VIB Center for Plant Systems Biology, VIB, Gent, Belgium
attributes.tag: INSDC first public
attributes.value: 2019-10-16T17:04:25Z
attributes.tag: INSDC last update
attributes.value: 2019-06-27T12:41:31Z
attributes.tag: INSDC status
attributes.value: public
attributes.tag: Submitter Id
attributes.value: E-MTAB-8095:exp2_FT_T_TP1_P1
attributes.tag: broker name
attributes.value: ArrayExpress
attributes.tag: cultivar
attributes.value: B104
attributes.tag: developmental stage
attributes.value: LP.04 four leaves visible stage
attributes.tag: genotype
attributes.value: GA2OX::PLA1
attributes.tag: growth condition
attributes.value: field study
attributes.tag: organism part
attributes.value: leaf base
attributes.tag: sample name
attributes.value: E-MTAB-8095:exp2_FT_T_TP1_P1
attributes.tag: sampling time point
attributes.value: 2 days after leaf 4 appearance
attributes.tag: scientific_name
attributes.value: Zea mays