|
Status |
Public on Mar 20, 2017 |
Title |
Biofilm 1 |
Sample type |
RNA |
|
|
Source name |
Porphyromonas gingivalis culture 96h
|
Organism |
Porphyromonas gingivalis |
Characteristics |
sampel type: biofilm
|
Treatment protocol |
Plates were incubated in anaerobic conditions at 37°C for 96 h
|
Growth protocol |
Planktonic cultures of P. gingivalis were grown anaerobically at 37ºC for 24 h in a protein-rich medium containing brain-heart infusion (BHI) A volume of 1.5 mL of P. gingivalis inoculums (concentration 10^8 CFU/mL) was placed in pre-sterilized polystyrene 24-well tissue culture plates (Greiner Bio-one) with or without the presence of sterile ceramic calcium hydroxyapatite discs
|
Extracted molecule |
total RNA |
Extraction protocol |
After 96 h of incubation, P. gingivalis planktonic and biofilm cells were harvested Total RNA was extracted from the harvested samples listed above, using the TRIzol® Max Bacterial RNA Isolation Kit (Ambion, Life Technologies, Carlsbad, CA, USA) RNA was quantified using a NanoDrop-1000 spectrophotometer and quality was monitored with the Agilent 2100 Bioanalyzer (Agilent Technologies, Santa Clara, CA).
|
Label |
Cy3
|
Label protocol |
Fluorescently labeled cDNA for microarray hybridizations was obtained by using the SuperScript Indirect cDNA Labeling System (Invitrogen)
|
|
|
Hybridization protocol |
Preparation of probes and hybridization was performed as described (One-Color Microarray Based Gene Expression Analysis Manual Ver. 6.5, Agilent Technologies) For each hybridization, 600 ng of Cy3 probes were mixed and added to 5 µL of 10x Blocking Agent and Nuclease free water in a 25 µL reaction Then, 25 µL from 2x GExHybridization buffer was added and mixed carefully The samples were placed on ice and quickly loaded onto arrays, hybridized at 65ºC for 17 h in a rotating Agilent hybridization oven After hybridization, microarrays were washed once in GE wash buffer 1 at room temperature (1 min) and once in GE Wash Buffer 2 at 37ºC (1 min).
|
Scan protocol |
Slides were scanned immediately after washing on the Agilent DNA Microarray Scanner using one color scan setting for 8x15K array slides
|
Description |
Gene expression after 96h Biofilm 1 sample 1 of 3
|
Data processing |
Images from Cy3 channel were equilibrated and captured with a high-resolution scanner (Agilent) and spots quantified using Feature Extraction software (Agilent) Background correction and normalization of data expression were performed using LIMMA
|
|
|
Submission date |
Mar 17, 2017 |
Last update date |
Mar 21, 2017 |
Contact name |
Patricia Romero Lastra |
E-mail(s) |
patrrome@ucm.es
|
Organization name |
Universidad Complutense de Madrid
|
Street address |
Plaza de Ramon y Cajal s/n
|
City |
Madrid |
ZIP/Postal code |
28040 |
Country |
Spain |
|
|
Platform ID |
GPL23193 |
Series (1) |
GSE96756 |
Comparative gene expression analysis of Porphyromonas gingivalis ATCC 33277 in planktonic and biofilm states |
|