title: Illumina HiSeq 2000 paired end sequencing; RNA-seq of Acidiphilium sp. C61 culture supplemented with 10 µM 2-Phenethylamine (PEA) against controls
design: RNA-seq of Acidiphilium sp. C61 culture supplemented with 10 µM 2-Phenethylamine (PEA) against controls
library_name: Sample 2_p
library_construction_protocol: Acidiphilium sp. C61 was isolated by Mori,J.F. and Lu,S. Acidiphilium sp. C61 cultures were cultivated in APPW+YE+Glucose medium with 0 µM or 10 µM PEA. APPW+ Yeast extract+ Glucose Harvested bacterial cells were lysed by TNS solution and subjected to beadbeating. Cell debris was separated by centrifugation and the obtained supernatant was first extracted with phenol-chloroform-isoamyl alcohol and second with chloroform-isoamyl alcohol. The obtained aqueous phase was subjected to overnight precipitation at 4°C with two volumes of polyethylene glycol 6000 and 5 μl glycogen. The resulting pellets were washed with ice-cold ethanol and suspended in 50 µl elution buffer. DNA were removed through DNAase digestion from nucleic acid and RNA was purified through Zymo RNA clean kit. Purified total RNA were used to do cDNA libraries. The library was constructed based on NEBNext ultra directional RNA library prep kit. Fragmented and primed RNA was reverse transcripted to first strand cDNA and further synthesized to second strand cDNA. End prep of cDNA library and  adaptor ligation were followed. Finallt, PCR library enrichment was conducted.
attributes.tag: Experimental Factor: compound
attributes.value: none