Short Name: TransientSPv11andSPv12-GCMS Assay Class: WET Assay Type: GCMS Title: Transient expression of constructs for SxPv1.1 and SxPv1.2 Description: The constructs for SxPv1.1 and SxPv1.2 were tested transiently in Nicotiana benthamiana leaves. Pheromone content was analysed via GC-MS pISA Assay creation date: 2020-07-23 pISA Assay creator: ElenaMG Lab manager: DO Sample collection protocol: Leaf samples were collected at 4 weeks after transplant for young stage and at 7 weeks for adult stage (early flowering). Samples were collected between 4 and 6 pm, frozen in liquid nitrogen immediately after collection, and ground afterwards. Extraction protocol:: 50mg of frozen, ground leaf samples were weighed in a 10mL headspace screw-cap vial and stabilized by adding 1mL of 5M CaCl2 and 150 μL of 500mM EDTA (pH = 7.5), after which they were sonicated for 5 minutes. Volatile compounds were captured by means of headspace solid phase microextraction (HS-SPME) with a 65 μm polydimethylsiloxane Chromatography protocol: Chromatography was performed on a DB5ms (60 m, 0.25 mm, 1 μm) capillary column (JandW) with helium as the carrier gas at a constant flow of 1.2mLxmin-1. For an initial identification of the pheromone peaks, oven programming conditions were 40°C for 2 min, 5°Cmin-1 ramp until reaching 280°C, and a final hold at 280°C for 5 min. Once the target peaks were identified, the oven conditions were changed to an initial temperature of 160°C for 2 min, 7°Cmin-1 ramp until 280°C, and a final hold at 280°C for 6 minutes to reduce the overall running time without losing resolution of the desired compounds Mass spectrometry protocol:: Electron impact ionization (EI), 70 eV ionization energy, MS source temperature 230ºC, MS quadrupole temperature 150ºC, single quadrupole detector, m Phenodata: ../../phenodata_20150819.txt Featuredata: Creation date: 2020-07-23 Extract ID: $_extr Extraction Method: HS-SPME Date Extraction: 2020-07-22 Derivatization or Labelling: none Date Derivatization or Labelling: 2020-07-22 Derivatized or labeled Extract ID: $_extrD Other Post Extraction Procedures: Storage: -80 degrees Date GC-MS Run: 2020-07-23 GC Instrument: 6890N gas chromatograph (Agilent Technologies) GC Autosampler Model: CombiPAL autosampler (CTC Analytics) GC Column model: DB5ms (60m, 0.25 mm, 1um) capillary column (JandW) GC Column type: capillary column Guard Column: MS Scan polarity: positive MS Scan mz range: 35-300 MS Instrument: 5975B mass spectrometer (Agilent Technologies) MS Ion source: electron ionization (EI) Mass analyzer: quadrupole mass filter Operator: RubenMF Notes: Data: https://doi.org/10.5281/zenodo.5810478

SEEK ID: https://fairdomhub.org/assays/1683

Experimental assay

Marko Petek

Projects: SUSPHIRE - Sustainable Bioproduction of Pheromones for Insect Pest Contr...

Investigation: _I_T21_SXPsysbio

Study: _S_P1_SPv1TransientExp

Assay position:

Assay type: Experimental Assay Type

Technology type: Technology Type

Organisms: No organisms