Difference between revisions of "Part:BBa K1412801:Experience"
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| − | 1. Transfer 50 μL bacterium solution (pLac-RBS (1.0)-cheZ-TT, pLac-RBS (0.01)-cheZ-TT, pLac-RBS (0.3)-cheZ-TT) into 5 ml new LB liquid medium whose chloromycetin concentration is 50ug/ml to culture for 12 hours in 37℃ horizontal rotators at 200 rpm. | + | 1. Transfer 50 μL bacterium solution (pLac-RBS(1.0)-<i>cheZ</i>-TT, pLac-RBS(0.01)-<i>cheZ</i>-TT, pLac-RBS(0.3)-<i>cheZ</i>-TT) into 5 ml new LB liquid medium whose chloromycetin concentration is 50ug/ml to culture for 12 hours in 37℃ horizontal rotators at 200 rpm. |
2. Then transfer another 50μL bacterium solution from the LB above to another new 5ml LB liquid medium whose chloromycetin concentration is 50ug/ml to culture in 37℃ horizontal rotators at 200rpm for 3 hours. | 2. Then transfer another 50μL bacterium solution from the LB above to another new 5ml LB liquid medium whose chloromycetin concentration is 50ug/ml to culture in 37℃ horizontal rotators at 200rpm for 3 hours. | ||
Latest revision as of 00:33, 18 October 2014
Protocol
Verification
Activiation
1. Transfer 50 μL bacterium solution (pLac-RBS(1.0)-cheZ-TT, pLac-RBS(0.01)-cheZ-TT, pLac-RBS(0.3)-cheZ-TT) into 5 ml new LB liquid medium whose chloromycetin concentration is 50ug/ml to culture for 12 hours in 37℃ horizontal rotators at 200 rpm.
2. Then transfer another 50μL bacterium solution from the LB above to another new 5ml LB liquid medium whose chloromycetin concentration is 50ug/ml to culture in 37℃ horizontal rotators at 200rpm for 3 hours.
Culture & Measurement
Culture
1. Firstly, draw three dots on a plate before using oily pen from the bottom of culture dish.
2. Secondly, stab 3μl bacterium medium into the M63 semisolid medium at the dots.
3. Finally, culture the bacteria in constant temperature and humidity incubator at 37℃ and observe the condition of bacterium growth.
Measurement
1. Observe the the condition of bacterium growth,and prepare a ruler.
2. When we can see a visible difference of the diameter of semi-solid medium with time pass by, begin to measure the diameter of bacterium colonies from the bottom of the semi-solid medium to avoiding the collapse of semi-solid medium.
3. Recording data: The diameter of initial colonies(0h) is recorded as R1. And the diameter of the colonies at 12h, 24h, 30h, 36h, 42h... as R2, R3, R4, R5, R6, R7….
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