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Revision as of 16:25, 13 August 2012 (view source) Konchan (Talk | contribs) (→Colony PCR) ← Older edit		Revision as of 23:51, 13 August 2012 (view source) Nili (Talk | contribs) Newer edit →
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	'''During the creation.'''<br>		'''During the creation.'''<br>
-	<big>'''Creating parts of Tar methylation region.'''<big>	+	'''Creating parts of Tar methylation region.'''
	==Date:8/9==		==Date:8/9==
	===Colony PCR===		===Colony PCR===

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Revision as of 23:51, 13 August 2012

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During the creation.
Creating parts of Tar methylation region.

Contents 1 Date:8/9 1.1 Colony PCR 2 Date:8/11 2.1 The purified DNA 2.2 PCR Production

Date:8/9

Colony PCR

Reagent

• TaKaRa Ex Taq(5units/μL) 0.5μL
• 10×Ex Taq buffer 10μL
• dNTP Mixture(2.5Meach) 8μL
• Primer F(10μM) 4μL
• Primer R(10μM) 4μL
• Template(E.coli DH5α)

→Reflection:We Took E.coli too many.You should take less E.coli.
Conditions of the thermal cycler

1. 95℃(5min)
2. 94℃(30sec)
3. 61℃(30sec)
4. 71℃(40sec)
5. 72℃(1min)
6. 4℃(Save)
   • 2~4:35cycle→Reflection:The number of cycles was less.So,We increased The number of cycles in 8/11.(50 cycles)

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Date:8/11

The purified DNA

1. Electrophoresis
   • Marker:pigment(buffer) 1μL,DNA molecule 2μL,TE buffer 3μL
   • Sample:pigment(buffer) 1μL,sample 5μL
   • Gel concentration:1.2%,Migration time:30min

→Reflection:Band was less.

1. Storage

PCR Production

1. Electrophoresis
   • The gel check and cut
2. DNA purification
3. Confirmation of electrophoresis
4. PCR
   • 50cycle
5. Storage

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