Samples are from the SRE4 experiment - an in situ experiment to determine fate and effects of different types of oils in the Antarctic marine environment. For details see: Powell S.M., Snape I., Bowman J.P., Thompson B.A.W., Stark J.S., McCammon S.A., Riddle M.J. 2005. A comparison of the short term effects of diesel fuel and lubricant oils on Antarctic benthic microbial communities. Journal ... of Experimental Marine Biology and Ecology 322:53-65.
Samples were analysed by denaturing gradient gel electrophoresis (DGGE) with primers specific for the Gammaproteobacteria.
Samples used were from Time2 (1 year)
Initial: T-1C; T-1E
SAB treatment: T2S
Primers: GAMFC: CGC CCG CCG CGC CCC GCG CCC GGC CCG CCG CCC CCG CCC GGG
TTA ATC GGA ATT ACT GG
GAMR: GGT AAG GTT CTT CGC GTT GCA T
50 ul (micro litre) reactions with HotStar (qiagen) mix, 5ul Q solution, 10 pmol (pico mol) each primer and 20 ng (nano gram) template DNA
cycling: 94C 15 minutes
35 cycles of:
94C 1 minutes
55C 1 minutes
72C 1 minutes
72C 20 minutes
DGGE was performed using D-Code system (BioRad).
Gel: 8% acryloamide, 30 - 65% denaturant with 2 cm stacking gel
1 x TAE, 60 degrees C, 80V 16 hours
Gel was pre-run for 20 minutes and lanes were flushed out after 15 minutes.
Gel was stained with Sybrgold.
Image captured using Storm Phosphorimager and ImageQuant v5.2 software (.gel files). The image files are called 151105#2.gel and 151105.tif
Band pattern results are in gammadgge.xls. The first column is the band position (or band name) and the remaining columns are samples with the first row being the sample name. The numbers indicates how many times the band appeared for that sample out of 2 DGGE runs.
This work was completed as part of ASAC projects 1228 and 2201 (ASAC_1228, ASAC_2201).