title: APS4_MI_2
organism: Auanema rhodensis
description: Protocols: For the mixed staged nematodes, M9 was gently added to 5 culture plates (6 cm) containing a healthy population of nematodes. We avoided disturbing the bacterial lawn and naturally let the nematodes start to swim in the buffer. This method reduced bacterial contamination in the samples during harvesting. These nematodes were then collected into a centrifuge tube washed twice with M9, and then frozen using liquid nitrogen. Tissues were homogenized for 1 min using a probe homogenizer. For RNA extraction, nematodes were first freeze-cracked in liquid nitrogen (2-3 times). Trizol was added to make up the volume to 500 µL and nematodes were shaken with a few sterile 0.5 mm glass beads on a BeadBeater homogenizer (20 s, 3 times with 30 s intervals). Subsequently, RNA was extracted using a standard chloroform approach and the pellet was dissolved in DEPC treated water and stored at -80 °C. Illumina TruSeq RNA sample Prep kit and protocol was used to construct the mRNA-seq libraries. Briefly, the poly-A containing RNA molecules were purified using poly-T oligo-attached magnetic beads. Following fragmentation of the mRNA, a first strand cDNA synthesis was performed using a reverse transcriptase and random primers, followed by a second strand cDNA synthesis. The cDNA fragments were then end repaired, an 'A' base was added, and finally the adapters were ligated. The resulting products were then purified twice and amplified by PCR to create the final cDNA library.
attributes.tag: ENA-FIRST-PUBLIC
attributes.value: 2019-02-12T17:04:20Z
attributes.tag: ENA-LAST-UPDATE
attributes.value: 2019-02-11T16:17:59Z
attributes.tag: External Id
attributes.value: SAMEA5332405
attributes.tag: INSDC center name
attributes.value: UNIVERSITY OF WARWICK
attributes.tag: INSDC first public
attributes.value: 2019-02-12T17:04:20Z
attributes.tag: INSDC last update
attributes.value: 2019-02-11T16:17:59Z
attributes.tag: INSDC status
attributes.value: public
attributes.tag: Submitter Id
attributes.value: E-MTAB-7667:APS4_MI_2
attributes.tag: age
attributes.value: mixed
attributes.tag: broker name
attributes.value: ArrayExpress
attributes.tag: developmental stage
attributes.value: mixed
attributes.tag: diet
attributes.value: OP50-1
attributes.tag: genotype
attributes.value: APS4
attributes.tag: individual
attributes.value: Manish Parihar
attributes.tag: organism part
attributes.value: whole organism
attributes.tag: replicate
attributes.value: 2
attributes.tag: sample name
attributes.value: E-MTAB-7667:APS4_MI_2
attributes.tag: scientific_name
attributes.value: Auanema rhodensis
attributes.tag: sex
attributes.value: mixed
attributes.tag: strain
attributes.value: APS4