title: clk-1 + 3
organism: Caenorhabditis elegans
description: Protocols: Worms of the following genotypes were used in this study: 1) clk-1(qm30) mutants, 2) clk-1(qm30); clk-1(nuc) mutants (rescued with a copy of clk-1 lacking the mitochondrial targeting signal and therefore localizing to the nucleus) and 3) clk-1(qm30); clk-1(WT) mutants (rescued with a wild-type copy of clk-1). The worms were cultured at 20 °C on nematode growth medium (NGM) agar plates seeded with OP50 strain Escherichia coli. Synchronized worms were harvested and snap frozen at L4 larval stage for total RNA isolation, or polysome profiling. For isolation of total RNA, worms were homogenized with a 5 mm steel bead using a TissueLyser II (Qiagen) for 5 min at frequency of 30 times/sec in the presence of TRIzol (Invitrogen) and isolation was continued according to manufacturer's protocol. Polysomal fractions from two experiments were pooled and mRNA was extracted using TRIzol LS (Invitrogen) according to the manufacturer's protocol. Contaminating genomic DNA was removed using RNase-Free DNase (Qiagen) and samples were cleaned up with the RNeasy MinElute Cleanup Kit (Qiagen). Samples were processed for Illumina using the NEBNext Ultra Directional RNA Library Prep Kit (NEB #E7420) according to manufacturer's instructions. Briefly, rRNA was depleted from total RNA using the rRNA depletion kit (NEB# E6310). After fragmentation of the rRNA reduced RNA, a cDNA synthesis was performed in order to ligate with the sequencing adapters and PCR amplification of the resulting product.
attributes.tag: ENA-FIRST-PUBLIC
attributes.value: 2018-08-28T17:03:21Z
attributes.tag: ENA-LAST-UPDATE
attributes.value: 2018-08-24T17:31:37Z
attributes.tag: External Id
attributes.value: SAMEA4858554
attributes.tag: INSDC center name
attributes.value: KU Leuven, Centre of Microbial and Plant Genetics (CMPG)
attributes.tag: INSDC first public
attributes.value: 2018-08-28T17:03:21Z
attributes.tag: INSDC last update
attributes.value: 2018-08-24T17:31:37Z
attributes.tag: INSDC status
attributes.value: public
attributes.tag: Submitter Id
attributes.value: E-MTAB-7166:clk-1 + 3
attributes.tag: broker name
attributes.value: ArrayExpress
attributes.tag: developmental stage
attributes.value: L4 larva
attributes.tag: fraction
attributes.value: total RNA
attributes.tag: genotype
attributes.value: clk-1(qm30)
attributes.tag: sample name
attributes.value: E-MTAB-7166:clk-1 + 3
attributes.tag: scientific_name
attributes.value: Caenorhabditis elegans
attributes.tag: strain
attributes.value: MQ130