Comment[GEAAccession]	E-GEAD-953
MAGE-TAB Version	1.1
Investigation Title	Gene regulated by luxS in Staphylococcus aureus (CBX75)
Experiment Description	Autoinducer 2 (AI-2) is synthesized by LuxS enzyme and has been suggested as a universal language for interspecies communication shared by Gram-negative and Gram-positive bacteria. The signaling functions of LuxS/AI-2 in Staphylococcus aureus, a ubiquitous and opportunistic pathogen causing nosocomial infections worldwide, remain obscure. Here we show that LuxS regulates a number of genes according to the transcriptional profiling in S. aureus strain NCTC8325 and that inactivation of luxS gene resulted in decreased susceptibility to cell-wall-synthesis inhibitor antibiotics, enhanced biofilm formation, decreased autolysis and up-regulated vancomycin-resistance associated VraSR two-component regulatory system. These alterations were restored by either genetic complementation or addition of exogenous AI-2, suggesting that the LuxS/AI-2 system in S. aureus is associated with these physiological functions mainly by quorum sensing. Our findings add new understanding to AI-2 quorum sensing regulation in S. aureus and provide novel clues for antimicrobial chemotherapy of Staphylococal infection.
Experimental Design	genetic modification design
Experimental Factor Name	sample_name
Experimental Factor Type	sample_name
Person Last Name	xue	Shang
Person First Name	Ting	Fei
Person Affiliation	University of Science and Technology of China	
Person Roles	submitter	
Public Release Date	2025-01-31
Protocol Name	P-GEAD-3914	P-GEAD-3915	P-GEAD-3916	P-GEAD-3917	P-GEAD-3918	P-GEAD-3919	P-GEAD-3920	P-GEAD-3921
Protocol Type	sample collection protocol	nucleic acid extraction protocol	nucleic acid labeling protocol	nucleic acid hybridization to array protocol	array scanning and feature extraction protocol	normalization data transformation protocol	growth protocol	treatment protocol
Protocol Description	none	TRIzol Reagent from Invitrogen Life Technologies has been used to isolate total RNA from Staphylococcus aureus. Follow the instructions provided by the supplier and, when necessary, use the steps outlined specifically for samples with high starch and/or high lipid content. QIAzol Lysis Reagent from QIAGEN can also be used.	Use GeneChipregistered mark DNA Labeling Reagent (Affymetrix, P/N 900542) to label the 3' termini of the fragmentation products. 1. Prepare the following reaction mix 2. Incubate the reaction at 37degree C for 60 minutes. 3. Stop the reaction by adding 2 microL of 0.5M EDTA. 4. The target is ready to be hybridized onto probe arrays, as described in Prokaryotic Target Hybridization on page 119. Alternatively, it may be stored at -20degree C for later use. To estimate the labeling efficiency, a gel-shift assay can be performed. In general, greater than 90% of the fragments should be labeled and, therefore, shifted.	Reagents and Materials Required The following reagents and materials have been tested and evaluated by Affymetrix scientists. Information and part numbers listed are based on U.S. catalog information. For supplier information, please refer to the Supplier Reference List in Appendix G of this manual. GeneChip Hybridization, Wash and Stain Kit: Affymetrix P/N 900720 (30 reactions) Hybridization Module, box 1 ? 2x Hybridization Mix ? DMSO ? Nuclease-free Water ? Control Oligo b2, 3 nM: Affymetrix, P/N 900301 Miscellaneous Supplies ? Hybridization Oven 640: Affymetrix, P/N 800138 (110V) or 800139 (220V) ? Sterile, RNase-free, microcentrifuge vials, 1.5 mL: USA Scientific,P/N 1415-2600 (or equivalent) ? Micropipettors, (P-2, P-20, P-200, P-1000): Rainin Pipetman (or equivalent) ? Sterile-barrier pipette tips and non-barrier pipette tips method: 1. Prepare the following hybridization cocktail. 2. Equilibrate probe array to room temperature immediately before use. 3. based on the format of the array type used, refer  to Table 6.2 to add the appropriate volume of hybridization cocktail. 4. Place probe array in the hybridization oven set at the temperatures indicated below. 5. To avoid stress to the motor, load probe arrays in a balanced configuration around axis. Rotate at 60  rpm. 6. Hybridize for 16 hours.During the latter part of the 16-hour hybridization, proceed to Chapter 7 to prepare reagents for the washing and staining steps required immediately after completion of  hybridization.	This protocol was provided by the Affymetrix., 1. Select Run -> Scanner from the menu bar. Alternatively, click the Start Scan icon in the tool bar. The Scanner dialog box appears with a drop-down list of experiments that have not been run. 2. Select the experiment name that corresponds to the probe array to be scanned. A previously run experiment can also be selected by using the Include Scanned Experiments option box. After selecting this option, previously scanned experiments appear in the drop-down list. 3. by default, for the GeneArrayregistered mark Scanner only, after selecting the experiment the number [2] is displayed in the Number of Scans box to perform the recommended 2x image scan. For the GeneChip? Scanner 3000, only one scan is required. 4. Once the experiment has been selected, click the Start button. A dialog box prompts you to load a sample into the scanner. 5. If you are using the GeneArrayregistered mark Scanner, click the Options button to check for the correct pixel value and wavelength of the laser beam. - Pixel value = 3 microm - Wavelength = 570 nm If you are using the GeneChip Scanner 3000, pixel resolution and wavelength are preset and cannot be changed. 6. Open the sample door on the scanner and insert the probe array into the holder. Do not force the probe array into the holder. Close the sample door of the scanner. If you are using the GeneChip Scanner 3000, do not attempt to close the door by hand. The door closes automatically through the User Interface when start scan is selected or the scanner goes into stand-by mode. 7. Click OK in the Start Scanner dialog box. The scanner begins scanning the probe array and acquiring data. When Scan in Progress is Selected from the View menu, the probe array image appears on the screen as the scan progresses.	Prior to conducting analysis of an array, the quality of the array image (.dat file) should be assessed following the guidelines found in the section \"Guidelines for Assessing Data Quality.\" When conducting a Comparison Analysis it is important to ensure that the scaling strategy used for the Comparison Analysis is the same as that used to generate the baseline file. To examine the analysis settings of the baseline file, right click the baseline .chp file in the Data File Tree and select \"Information.\", See \"http://www.affymetrix.com/support/downloads/manuals/data_analysis_fundamentals_manual.pdf\"., A 2-fold induction ratio as a cutoff limit was used by us to compare the transcription profile between the luxS mutant and the wild type strain.	The culture condition of the strain: S. aureus and its derivative strains were grown in TSb (tryptic soy broth, soybean-casein digest medium USP; Oxoid) medium, and when necessary, erythromycin (2.5 mg/l), and chloramphenicol (15 mg/l) were supplemented.	The cultures were incubated at 37degree C with shaking at 250 rpm in 50-ml flasks. When the cultures reached the appropriate OD600 = 1.7, the cells were harvested for RNA extraction.
SDRF File	E-GEAD-953.sdrf.txt
Comment[Number of channel]	single-channel
Comment[Array Design REF]	A-GEAD-158
Comment[AEExperimentType]	transcription profiling by array
Comment[SecondaryAccession]	CBX75
Comment[BioProject]	PRJDB20052
Comment[CIBEX Accept Date]	2010-01-01
Comment[Last Update Date]	2025-01-31