Brassicogethes aeneus

Adult Meligethes aeneus shotgun transcriptome

mRNA was purified from 1µg total RNA by exonuclease digestion followed by LiCl precipitation (mRNA-Only Eucaryotic mRNA Isolation Kit, Epicentre, Madison, WI, USA); 1µg mRNA was used for first-strand cDNA synthesis. cDNA synthesis and amplification was done according to the Mint-Universal cDNA Synthesis Kit user manual (Evrogen, Moscow, Russia ); 800 ng amplified cDNA were used as starting material in the normalization reaction using the Trimmer Kit (Evrogen, Moscow, Russia), normalized material was re-amplified for 18 cycles; 2 µg of normalized cDNA was digested with 10 Units SfiI for 2hours at 48°C. Fragments larger than 800b p were isolated from a LMP Agarose Gel and purified using the MinElute Gel Extraction Kit (Qiagen, Hilden, Germany). 200ng purified cDNA fragments were ligated to 100ng Sfi cut and dephosphorylated pDNR-lib Vector (Clontech) in 10µl Volume using the Fast Ligation Kit (NEB ,Ipswich, MA,USA). Ligations were desalted by Ethanol precipitation, and re-dissolved in 10µl water. 3 times 1,5 µl desalted ligation was used to transform NEB10b competent cells (NEB, Ipswich, MA,USA).; Library generation for the 454 FLX sequencing was carried out according to the manufacturer’s standard protocols (Roche/454 life sciences, Branford, CT 06405, USA).; In short, the concatenated inserts were sheared randomly by nebulization to fragments ranging in size from 400 bp to 900 bp. These fragments were end polished and the 454 A and B adaptors that are required for the emulsion PCR and sequencing were added to the the ends of the fragments by ligation. The resulting fragment library was sequenced on 1/2 picotiterplate (PTP) on the GS FLX using the Roche/454 FLX+ chemistry. A total of 554 020sequence reads with an average read length of 505 nucleotides were obtained.

{'label': 'adult', 'body_site': 'Body of invertebrate', 'geo_loc_name': 'Poland', 'specific_host': 'Brassica napus', 'collection_date': '2012-04', 'isolation_source': 'adult beetles'}

{'dev stage': 'Adult', 'treatment': 'Control', 'external id': 'SAMEA2063032', 'sample name': 'Maen AD-Control', 'insdc status': 'public', 'submitter id': 'Maen AD-Control', 'ena-last-update': '2018-03-09T10:12:14Z', 'scientific_name': 'Brassicogethes aeneus', 'ena-first-public': '2014-01-09T17:03:38Z', 'insdc center name': 'MPICE', 'insdc last update': '2018-03-09T10:12:14Z', 'insdc first public': '2014-01-09T17:03:38Z'}

{'dev stage': 'Adult', 'treatment': 'Immune-Induced', 'external id': 'SAMEA2063033', 'sample name': 'Maen AD-Immune', 'insdc status': 'public', 'submitter id': 'Maen AD-Immune', 'ena-last-update': '2018-03-09T10:12:14Z', 'scientific_name': 'Brassicogethes aeneus', 'ena-first-public': '2014-01-09T17:03:38Z', 'insdc center name': 'MPICE', 'insdc last update': '2018-03-09T10:12:14Z', 'insdc first public': '2014-01-09T17:03:38Z'}