Quorum+Sensing+Project

http://www.ncbi.nlm.nih.gov/pmc/articles/PMC205467/pdf/jbacter00028-0314.pdf

Use fishtank air pump and bubble stone to aerate? requires something bigger than culture tubes http://www.amazon.com/Elite-800-Air-Pump-A800/dp/B0002AQFZ2


 * Inhibition methods:
 * In vitro production of AHL blockage:
 * L/D-S-adenosylhomocysteine
 * sinefungin
 * butyryl-S-adenosylmethionine (butyryl-SAM)
 * Increase pH to above 7, causing lactonolysis (ring opening of AHL)
 * Temperature increase?
 * Enzymes:?
 * Hypobromous and hypochlorous acids
 * Analogue of AHL molecule:
 * Natural:
 * crown vetch
 * carrot
 * soybean
 * tomato
 * habanero
 * garlic


 * Photobacterium broth:
 * http://www.fotodyne.com/content/epdKC_strains (buying, $20 per liter, also stocks photobacterial agar plates and vibrio fischeri)
 * http://www.carolina.com/product/living+organisms/biological+media/dehydrated+media/phenol+red+broth+base%2C+dehydrated+medium%2C+500+g.do?keyword=photobacterium+broth&sortby=bestMatches&page=1 Liquid media
 * http://www.carolina.com/product/living+organisms/biological+media/prepared+media+bottles+and+tubes/peptone+iron+agar%2C+prepared+media+tubes%2C+pack+10.do?keyword=photobacterium&sortby=bestMatches&page=1 Prepared media tubes
 * Ways to count bacteria:
 * http://www.textbookofbacteriology.net/growth.html
 * http://www.cellprofiler.org/index.shtml (free colony counting software)
 * http://www.freescienceproject.com/A/projects/BIO03.htm
 * Serial dilutions from liquid cultures:
 * http://www.disknet.com/indiana_biolab/b038.htm
 * http://www2.muw.edu/~lbrandon/Micro/enumeration.doc
 * http://www.waksmanfoundation.org/labs/rochester/dilution.htm
 * http://www.waksmanfoundation.org/labs/rochester/food.htm


 * VAI1
 * http://www.nottingham.ac.uk/quorum/compounds.htm
 * http://www.sigmaaldrich.com/catalog/ProductDetail.do?N4=K3255|FLUKA&N5=SEARCH_CONCAT_PNO|BRAND_KEY&F=SPEC&lang=en_US

http://rstb.royalsocietypublishing.org/content/362/1483.toc (more quorum sensing papers)

TECHNIQUES http://www.sciencebuddies.org/science-fair-projects/project_ideas/MicroBio_Techniques.shtml http://www.protocol-online.org/biology-forums/posts/23590more1.html http://www.sciencebuddies.org/science-fair-projects/project_ideas/MicroBio_Storing_Bacterial_Strains.shtml

protocols: http://2010.igem.org/Team:Michigan/Project#Hy-Bi:_Quorum_Sensing

===** [|Using Bioluminescent Bacteria to Detect Water Contaminants] (details method of "measuring" bioluminescence and other procedures) ** === (Supplies in this experiment obtained from http://www.carolina.com/)

http://www.pleasanton.k12.ca.us/avhsweb/thiel/creek/research/00/shelley_jasper/procedure.html

Control Garlic pH above 7 Analogue: decanoyl homoserine lactone (http://www.sigmaaldrich.com/catalog/ProductDetail.do?lang=en&N4=17248|SIGMA&N5=SEARCH_CONCAT_PNO|BRAND_KEY&F=SPEC) (http://www.caymanchem.com/app/template/Product.vm/catalog/10011201/a/z;jsessionid=091F728FDD78495DB72E666E835320DE)

Do garlic and pH variables first, then if time attempt analogue.

TO DO:


 * Imaging:
 * Buy external webcam
 * Build a lightproof box
 * http://gawker.sourceforge.net/Gawker.html to create timelapse movies
 * Vernier light sensor available
 * Things to buy:
 * Photobacterium broth (http://www.carolina.com/product/living+organisms/biological+media/dehydrated+media/phenol+red+broth+base%2C+dehydrated+medium%2C+500+g.do?keyword=photobacterium&sortby=bestMatches&page=1) 1 100g bottle at $44
 * Photobacterium agar, ~50 dishes (http://www.carolina.com/product/living+organisms/biological+media/nutrient+agar%2C+prepared+media+bottle%2C+125+ml.do?keyword=photobacterium&sortby=bestMatches) 3 bottles @$6.50 each
 * Filters: http://www.fishersci.com/wps/portal/PRODUCTDETAIL?prodcutdetail='prod'&productId=606447&catalogId=29103&matchedCatNo=0979012C||0979012D||0979012E||0979012F||0979012G&pos=7&catCode=HC_SC&endecaSearchQuery=%23store%3DHealthCare%23N%3D0%23rpp%3D15&fromCat=yes&keepSessionSearchOutPut=true&fromSearch=Y&searchKey=filter||filtrations||filtration||filtering||paper||filters||papers&highlightProductsItemsFlag=Y
 * 1 pack of 100 for $11.64
 * Vibrio fischeri (http://www.carolina.com/product/vibrio+anguillarum%2C+living%2C+tube.do?keyword=vibrio+fischeri&sortby=bestMatches) $9.50
 * Culture tubes (http://www.sigmaaldrich.com/catalog/ProductDetail.do?lang=en&N4=Z376787|SIGMA&N5=SEARCH_CONCAT_PNO|BRAND_KEY&F=SPEC) 1000 tubes @ $34
 * Petri dishes, ~50 (http://www.amazon.com/Pack-Sterile-Plastic-Petri-Dishes/dp/B001S4D36A/), $7.50 for 20

$44+19.5+11.64+9.5+34+22.5= $141.14


 * Questions:
 * Culture tubes necessary or autoclave flasks?

Do garlic and pH variables first to become accustomed to 1. Grow control (both liquid and agar) of untreated V. fischeri, measure light output in lightproof box 2. Determine required concentration of bacteria to glow (optional) 3. Determine time required for V. fischeri to reach adequate concentrations 4. In basic media and agar, grow V. fischeri and measure light output after time in 3 (use timelapse camera?) 5.

In individuals with cystic fibriosis, the bacteria Pseudomonas aeruginosa can wreak havoc on the lungs. The major reason that P. aeruginosa is so harmful is due to its ability to form a biofilm, or a sheetlike formation of adherent cells. The ability of P. aeruginosa to form a biofilm is regulated by quorum sensing, a process that allows bacteria to communicate and act as a multicellular organism. Vibrio fischeri, a harmless bioluminescent bacteria, uses a similar quorum sensing mechanism to regulate bioluminescence. I plan to use Vibrio fischeri as a model to explore various methods of interfering with the process of quorum sensing that can hopefully be used to treat patients infected by P. aeruginosa.