PGLO transformation

PGLO transformation

1. Label two B12 competant (able to take DNA) Ecoli cell tubes and (-) PGLO, (+) PGLO and place on ice.
2. Transfer 250 µm of cold transformation solution CaCl2 into each tube
3. Place tubes on crushed ice
4. Using pipet 10 µL of PGLO plamid (DNA) into + PLGO tube.
5. Incubate tubes onn ice for 10 mins
6. Lable 4 agar plates on the bottom 
(1) LB with (-) PGLO
(2) LB/Amplara with (-) PGLO
(3) LB/Amplara (+) PGLO
(4) LB/Amplara/Ara  (+) PGLO

Note: We must test the result that
(1) Cell can grow with lots of growth
(2) No growth
(3) Growth not much
(4) Growth/ Glow (Has a light)





7. Heat shock using both for  (-) PGLO, (+) PGLO tubes into water bath at 42  degrees celsius for 50 sec. Then place back on ice in 2 mins.
8. Remove from ice and rack. Add 250 µm LB broth into tubes and incubate at room temp for 10 min
9. Grenly flick closed tubes to mix pipet 100  µm from each tube to corresponding plates 
 - Titrate Lb/ Amphata plates
 - Other plates 50 µm 
10. Spread evenly on plates
11. Stach upside down and tape tgether. Place in incubator in 37 degrees celsius  until next day.


(-) control cell dont have plow, no contamination
(+) control cell have plow, has contamination

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