Team:Chiba/jk/γ/Trl

(Difference between revisions)

Revision as of 23:52, 29 October 2008

インダクションをかけてからいち早く、確認できる出力を見つけること
To find the earliest output gene after induction

shaking incubator

Innova 4200 Benchtop or Floor-Stackable Incubator Shaker(37°C)

46-well plate(deep well)

96-well plate(deep well)

Fluoroskan Ascent 2.5(program:Ascent Software Version 2.6)

Beckman Allegratm X-12R Centrifuga(Beckman Coulter)

試薬(reagents)

Fig. レポーターのタイムレスポンスの実験方法

Strain:XL10G Kan^R

Pre-culture
1. Picked and cultured the following plate in 2mL of LB:
  1. LB-Amp+0.2 % Glucose, (pGFPuv, pLac-Venus YFP, pLac-mCherry, pUC19)
  2. LB-Amp, ([http://partsregistry.org/Part:BBa_T9002 BBa_T9002], [http://partsregistry.org/Part:BBa_K084003 BBa_K084003])
2. Cultured at 37°C for 12h.
Culture
1. Dilute pre-cultures and add to new LB medium.
  1. LB-Amp+0.2 % Glucose, (pGFPuv, pLac-Venus YFP, pLac-mCherry, pUC19)
  2. LB-Amp, ([http://partsregistry.org/Part:BBa_T9002 BBa_T9002], [http://partsregistry.org/Part:BBa_K084003 BBa_K084003])
2. Cultured at 37°C for about 6 h
Wash
1. Transfer 10mL each of the culture to 50mL centrifuge tubes.
2. Centrifuged for 6min at 3600rpm,20°C and discarded the supernatant.
3. Add physiological saline and resuspention
4. Repeated wash twice.
5. Add M9 minimal medium.
Mix
1. Dispens each culture into 48-well deep well or 96-well deep well
2. Add IPTG fainal conc. 0.2 mM (pGFPuv, pLac-Venus YFP, pLac-mCherry, pUC19)
3. Add AHL fainal conc. 100nM ([http://partsregistry.org/Part:BBa_T9002 BBa_T9002], [http://partsregistry.org/Part:BBa_K084003 BBa_K084003])
Measure fluorescence intensity every 1 h.

Result

ホーム	メンバー紹介	プロジェクト紹介	Parts Submitted to the Registry	モデリング	ノート

@@ Line 9: / Line 9: @@
 ===装置&試薬(apparatus&reagents)===
-'''装置（apparatus)'''
+*Equipment
-*しんとう培養器(shaking incubator)(37℃,30℃)
+:shaking incubator
-*48 well plate(deep well)
+::Innova 4200 Benchtop or Floor-Stackable Incubator Shaker(37&deg;C)
-*Fluoroskan Ascent 2.5(program:Ascent Software Version 2.6)
+:46-well plate(deep well)
+:96-well plate(deep well)
+:Fluoroskan Ascent 2.5(program:Ascent Software Version 2.6)
+:Beckman Allegratm X-12R Centrifuga(Beckman Coulter)
 '''試薬(reagents)'''
 *AHL(100uM,5uM,100nM)