From 2010.igem.org
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| - | ==2010/08/26(Bambi75)==
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| - | ==Make Plates==
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| - | ===member===
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| - | NEX , Bambi75 and watachin
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| - |
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| - | ===Materials===
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| - | *RO water 200ml
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| - | *LB Broth 4g
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| - | *Cam(50μg/L) 10ml
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| - |
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| - | ===Procedure===
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| - | ① mix materials.
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| - |
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| - | ② Divide ①equally and make 10 plates.
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| - |
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| - | ==Separation of A.xylinum==
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| - |
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| - | ===member===
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| - | easily and naoto
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| - |
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| - | ===Materials===
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| - | *the plate(made on August 25th).
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| - |
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| - | ===Procedure===
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| - | ①extract material 2ml.
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| - |
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| - | ②centrifuge ① 10000rpm/5min.
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| - |
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| - | ==PCR==
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| - |
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| - | ===member===
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| - | same above
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| - |
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| - | ===Materials===
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| - | *2×PCR buffer 25×4μl
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| - | *2mM dNTP 10×4μl
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| - | *10mM primer(sense)bcsA,B,C and D 2.5μl each
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| - | *10mM primer(antisense)bcsA,B,C and D 2.5μl each
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| - | *template DNA a little
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| - | *Q water 9×4μl
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| - | *KOD FX 0.5×4μl
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| - |
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| - | ===Procedure===
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| - | ①mix all materials for 4 tubes.
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| - |
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| - | ②elongation
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| - | *bcsA,bcsB and bcsC
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| - | **94℃ 2min
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| - | **98℃ 10sec☆
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| - | **55℃ 30sec
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| - | **68℃ 4min★
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| - | **68℃ 7min
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| - | **10℃ ∞
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| - | *bcsD
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| - | **94℃ 2min
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| - | **98℃ 10sec☆
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| - | **55℃ 30sec
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| - | **68℃ 1min★
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| - | **68℃ 7min
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| - | **10℃ ∞
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| - | ※30cycle ☆ to ★.
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| - |
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| - | ==PCR==
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| - |
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| - | ===member===
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| - | NEX , Bambi75 and watachin
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| - |
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| - | ===Materials===
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| - | *sterilized water 71μl
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| - | *Ex taq buffer 10μl
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| - | *dNTP mix 8μl
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| - | *Ex taq 1μl
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| - | *primer(bcsC sense/antisense) 5μl each
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| - |
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| - | ===Procedure===
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| - | ①mix materials
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| - |
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| - | ②divide ① into 2 tubes.
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| - |
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| - | ③elongation
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| - | **95℃ 3min
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| - | **96℃ 1min☆
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| - | **55℃ 7min
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| - | **72℃ 1min★
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| - | **10℃ ∞
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| - | ※30 cycle ☆to★.
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Latest revision as of 06:08, 8 September 2010
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