characteristics.text: ERC000011
characteristics.characteristic: ENA-CHECKLIST
characteristics.text: 2015-04-01T17:03:47Z
characteristics.characteristic: ENA-FIRST-PUBLIC
characteristics.text: 2018-03-08T17:29:03Z
characteristics.characteristic: ENA-LAST-UPDATE
characteristics.text: SAMEA2479553
characteristics.characteristic: External Id
characteristics.text: The Pirbright Institute
characteristics.characteristic: INSDC center name
characteristics.text: 2015-04-01T17:03:47Z
characteristics.characteristic: INSDC first public
characteristics.text: 2018-03-08T17:29:03Z
characteristics.characteristic: INSDC last update
characteristics.text: public
characteristics.characteristic: INSDC status
characteristics.text: ERS445038
characteristics.characteristic: SRA accession
characteristics.text: E-MTAB-2527:L1_12h_M
characteristics.characteristic: Submitter Id
characteristics.text: 12 hour
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/UO_0000032
characteristics.characteristic: age
characteristics.text: ArrayExpress
characteristics.ontologyTerms: http://www.ebi.ac.uk/efo/EFO_0009737
characteristics.characteristic: broker name
characteristics.text: African malaria mosquito
characteristics.characteristic: common name
characteristics.text: Protocols: The mosquitoes were kept in the insectary conditions at 27C, 80% RH, and 12h light/12h dark cycle. Larvae were reared in pans with de-ionized water and fed daily with ground fish food (Tetramin). Pans containing mature 4th instar larvae were inspected every hour, and newly pupated individuals transferred into marked pans with clean de-ionized water. Samples were collected at appropriate time intervals and sexed prior to grinding in TRIzol. The samples were ground in TRIzol and stored at -70C. Total RNA was isolated using PureLink RNA Mini Kit (Ambion) according to the manufacturer's protocol. RNA quality was evaluated using Qubit RNA assay (Life Technologies) and on a PerkinElmer GX using the HT RNA Reagent Kit (PerkinElmer) prior to construction of libraries. The libraries were constructed with the PerkinElmer Sciclone NGS Workstation using the TruSeq RNA protocol (Illumina Inc.). mRNA was isolated from 1 ug of total RNA by a poly-A+ pull down using biotin beads and fragmented. First strand cDNA was synthesised, followed by the second strand synthesis. The ends of the samples were repaired using the 3' to 5' exonuclease activity to remove the 3' overhangs, and the polymerase activity to fill in the 5' overhangs, creating blunt ends. A single 'A' nucleotide was added to the 3' ends of the blunt fragments to prevent them from ligating to one another during the adapter ligation reaction. A corresponding single 'T' nucleotide on the 3' end of the adapter provided a complementary overhang for ligating the adapter to the fragment. This strategy ensured a low rate of chimera formation. The ligated products were subjected to a bead-based size selection using Beckman Coulter XP beads (Beckman Coulter). This removed the majority of un-ligated adapters, as well as any adapters that may have ligated to one another. Prior to hybridisation to the flow cell, the samples were amplified by PCR to selectively enrich fragments with adapter molecules on both ends. The PCR was performed with a PCR primer cocktail that annealed to the ends of the adapters. The insert size of the libraries was verified by running an aliquot of the DNA library on a PerkinElmer GX using the High Sensitivity DNA chip (PerkinElmer) and the concentration was determined by using a High Sensitivity Qubit assay (Life Technologies) and q-PCR. Prior to hybridisation to the flow cell, the samples were amplified by PCR to selectively enrich fragments with adapter molecules on both ends. The PCR was performed with a PCR primer cocktail that annealed to the ends of the adapter. The insert size of the libraries was verified by running an aliquot of the DNA library on a PerkinElmer GX using the High Sensitivity DNA chip (PerkinElmer) and the concentration was determined by using a High Sensitivity Qubit assay (Life Technologies) and q-PCR.
characteristics.characteristic: description
characteristics.text: first instar larva
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/FBdv_00005337
characteristics.characteristic: developmental stage
characteristics.text: Anopheles gambiae
characteristics.characteristic: organism
characteristics.text: Anopheles gambiae
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/NCBITaxon_7165
characteristics.characteristic: organism
characteristics.text: Anopheles gambiae
characteristics.characteristic: scientific_name
characteristics.text: male
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/PATO_0000384
characteristics.characteristic: sex
characteristics.text: fresh specimen
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/OBI_0000971
characteristics.characteristic: specimen with known storage state
characteristics.text: Pimperena
characteristics.characteristic: strain
characteristics.text: L1_12h_M
characteristics.characteristic: title