Procedureee Flashcards
- Prepare the needed materials (_______prior to experiment to ensure sterility).
- Prepare the PEG-200 solution by weighing__________ and mix with_______ to achieve a _____ratio solution.
Heat the mixture to completely dissolve the PEG-200.
autoclave
2.5g of PEG-200
20mL sterile nuclease-free water
1:8
- Prepare the NaOH solution by weighing ________and mix it with________ to achieve a_____ solution. Mix gently.
1.6g of NaOH
20 mL of nuclease-free water
clear
- Combine the (2)
PEG-200 solution and NaOH solution
Volume (uL) needed per bacterial sample
NaOH 2M
5 uL
Volume (uL) needed per bacterial sample
PEG 400
60 uL
Volume (uL) needed per bacterial sample
Sterile Nuclease-Free Water
25 uL
PEG NAOH
*This solution is stable for_____ when stored at _____in a_____.
1 month
room tenperature
glass bottle
- Obtain a bacterial sample.
This sample may be an overnight culture of a bacteria or yeast in_____, or bacterial or yeast colonies suspende in_____,______ or_____.
(3) may also be used.
nutrient broth
water, nutrient broth, or saline
Urine, cerebrospinal fluid, or plasma
PEG NAOH
*This solution is stable for_____ when stored at _____in a_____.
1 month
room tenperature
glass bottle
Tissue culture cells from one 24-well tissue culture dish scraped into ___________ with _________ may also be used to extract cellular DNA or viral DNA.
100 uL of DMEM (Dulbecco’s Modified Eagle Medium ) with
10% serum
If the source of bacterial sample is from agar plate, please do the following:
a) Obtain a loopful of_______ from an overnight agar plate culture using a______ or _____
b) Suspend the looplul colony in ________in a ______tube.
single bacterial colony
sterile inoculating loop or a sterile applicator stick.
100 uL of sterile molecular grade water
1.5mL microcentrifuge
If the source of bacterial sample is from broth, please do the following:
a) Obtain ______of broth culture and place it in a_______
100 uL
1.5 mL microcentrifuge tube
- Prepare a______ set to _____
- Mix the samples using a_____.
- Centrifuge at _____ for ______.
- Remove _____of the____, leaving the bacterial sample suspended in the remaining_____.
- Add ______and mix briefly.
- Heat the mixture at _____ for ______in the water bath.
- Store the sample in the_____.
water bath set to 80°C.
vortex
6,082 x g for 4 min
90 uL ; superatant; 10 uL
90 uL of PEG-NaOH solution
80°C for 10 minutes
freezer
______the sample briefly before use in PCR.
Use_____ of the sample directly for a 15-uL total reaction volume in PCR.
Shake
2 uL
Final concentration (90 uL PEG-NaOH + 10 uL Bacteria)
PEG-NaOH
Bacterial sample
Volume
90
10
Concentration
100 Mm
60%
