Bacterial media
Bacterial media kdorfman Thu, 01/17/2013 - 18:53Freezing bacteria
Freezing bacteria kdorfman Sat, 12/05/2015 - 18:44To Freeze:
Pick a new colony, grow overnight in LB
inoculate LB from the overnight culture
Put 0.15 mL glycerol into sterile cryovial.
Add 0.85 mL mid-log culture (OD =~0.4). Pipette up and down to mix thoroughly
Freeze. Store in -80 if possible.
To thaw:
Scrape surface of frozen stock with sterile stick
Streak on agar plate
LB Broth
LB Broth kdorfman Fri, 09/25/2020 - 17:54LB broth from mix
LB broth from mix kdorfman Tue, 05/28/2013 - 17:48LB Broth Miller Luria-Bertani
Fisher DF0446-17-3 500 g $40.61.
25 g/L
LB from scratch
LB from scratch kdorfman Fri, 09/25/2020 - 17:38LB (Miller) from scratch
LB agar
LB agar kdorfman Thu, 01/17/2013 - 18:55LB agar granules
LB agar granules kdorfman Tue, 06/25/2019 - 20:01Make with granulated LB (Miller) agar, Fisher BP9724
LB agar powder
LB agar powder kdorfman Tue, 06/25/2019 - 20:00Make with LB (Miller) Agar powder, Difco 244510
LB broth + agar
LB broth + agar kdorfman Tue, 06/25/2019 - 20:0025 g LB + 15 g agar /liter
LB antibiotic
LB antibiotic kdorfman Wed, 10/01/2014 - 16:34Table of antibiotic stability in poured agar
line | antibiotic |
---|---|
black | Carbenicillin (ampicillin) |
green | IPTG |
LB-sugars
LB-sugars kdorfman Wed, 10/01/2014 - 16:19LB + sugars for Lac-operon work
sugar (or analog) | MW |
---|---|
mono saccharide | 180.2 |
disaccharide | 342.3 |
IPTG | 238.3 |
glucose, galactose, or fructose
Lactose is not very soluble in cold water.. Boil water, then set it to stir on a stir plate. Gradually tap in the lactose powder.
lactose, maltose, sucrose
IPTG
Filter sterilize
Add 50 µg/mL Kanamycin as indicated here.
MacConkey agar
MacConkey agar kdorfman Thu, 01/17/2013 - 19:41MacConkey Agar
- Fisher 212122 2 kg (Difco)
- Krackeler 10-211387 500g (via Sigma)
- HiMedia MacConkey agar, granulated GM081 (Fisher NC1876763, VWR 95020-204)
to distinguish Lac+ and Lac- bacterial strains.
50 g/L
(If you are going to make more than one bottle, turn on 65C waterbath in tissue culture room)
Turn on the stir-plate heat to 105C.
Withhold 50 mL of water to rinse the mouth of the bottle.
Heat the rest of the water to boiling in the bottle in the microwave.
Measure out the agar mix while the water is heating
Add the stir bar (CAREFULLY!) to the bottle of hot water
Put the bottle on the stir-plate
Add the agar mix to the bottle
Use the reserved water to rinse the granules stuck in the neck down to the liquid.
Stir and heat until granules are completely dissolved.
Cap loosely, wrap with foil, and autoclave.
(If you make more than one bottle, the extras go in the water bath till you are ready to pour)
leave stir bar in
put bottles on stir plate near sterile hood until handle-able
If you accidentally buy
MacConkey + bile+ CV
MacConkey + bile+ CV kdorfman Mon, 01/08/2024 - 16:00If you accidentally buy MacConkey without
- bile salts (1.5 g/L)
- Crystal violet (0.001g/L)
- NaCl (5 g/L)
Min med for GFP cells
Min med for GFP cells kdorfman Tue, 10/23/2018 - 15:53From https://wahoo.cns.umass.edu/node/857
For SfGFP cells, try M9 + 0.2% glycerol + 0.01 mM FeSO4
Mueller Hinton agar
Mueller Hinton agar kdorfman Thu, 01/17/2013 - 20:0590922 Mueller Hinton Broth 2 from Sigma
22 grams per liter, consisting of:
Casein acid hydrolysate 17.5
Beef extract 3.0
Starch 1.5
Final pH 7.3 +/- 0.2 at 25°C
P-Glo Bacterial media
P-Glo Bacterial media kdorfman Mon, 08/26/2019 - 22:26Make 60 mm plates
plain LB plates
LB/Amp (or Carb) plates (1 black stripe)
LB/Amp/Ara plates (1 black, 1 red stripe)
2019 - for QBoC:
- transformed pglo plasmid (1660405EDU) from the BioRad kit into
- DH5 alpha E. coli (Invitrogen 18265-017) from the -80
- Lyophilized cells (BioRad 1660408EDU) did not arrive in time
Used the Invitrogen protocol.
Plated on:
- LB
- LB amp
- LB amp ara
Saw gfp colonies on the arabinose plates. Collected, grew up in LB amp, froze in 15% glycerol.
LB-Amp-Ara
LB-Amp-Ara kdorfman Mon, 08/26/2019 - 22:27LB with Ampicilin or Carbenicilin and arabinose
Arabinose stock 200mg/mL in water, sterilized by filtration (=100 X)
Final concentration = 2 mg/mL
- Arabinose MW = 150.13
- 2 mg/mL = 2 g/L x 1 mol/150.13 g = 0.013M = 13 mM
Carbenicilin stock is 1000X
minimal medium (glycerol)
minimal medium (glycerol) bcrcstaff Wed, 11/14/2018 - 17:34For motility strains