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Revision as of 04:11, 27 October 2007 (view source) Hiroki (Talk | contribs) (→FliC-His generator) ← Older edit		Revision as of 04:14, 27 October 2007 (view source) Umeno (Talk | contribs) (→Experiment) Newer edit →
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	===[[Chiba/Flagella/FliC-His_Biobrick|FliC-his biobrick]]===		===[[Chiba/Flagella/FliC-His_Biobrick|FliC-his biobrick]]===
	====Experiment====		====Experiment====
-	*ここの文章の意味が伝わりにくいです。細かい話になりすぎるのであれば、この箇所はカットしてもいいと思います。byとよたろ	+	*Making Biobrick version of Flic-His and other stickly-FliCs. unfortunately, FliC has many EcoRI,SpeI,PstI sites. We have to eliminate them one by one by site-directed mutagenesis.
-	*Change puc19 vector into biobrick's vector.	+
-	(puc19 vectorの乗っているのでbiobrickのベクターに乗せる。)	+
-		+
-	*Broken restriction sites of enzyme(EcoRI,SpeI,PstI).	+
-	(制限酵素サイト(EcoRI,SpeI,PstI)をつぶす。)	+
-		+
-	*Can't insert vector directly,because FliC-His contains EcoRI,SpeI,PstI.	+
-	#Insert blunt end on one side and ApaI on the other side.	+
-	#Similarly do PCR and ligation in the vector side.	+
-		+
-	(FliC His-TagにはEcoRI,SpeI,PstIが含まれているために、そのままではvectorに入れられない。	+
-	#片側をblunt end もう一方をApaIの制限酵素サイトをつけ、PCRする。	+
-	#vector側も同様にPCRしLigationさせる。)	+
	====Results====		====Results====
	Not yet (as of 10/26/2007). Cloning is still underway -->		Not yet (as of 10/26/2007). Cloning is still underway -->

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Revision as of 04:14, 27 October 2007

Introduction | Project Design ( 1.Affinity Tag | 2.Communication Module | 3.Size Control ) | Making Marimos | Our Goal Acknowledgements | Team Members | [http://chem.tf.chiba-u.jp/igem/ iGEM Chiba Website] | メンバ連絡簿

Making Marimos

Parts Construction

Because we found that FliC code includes restriction enzyme(EcoRI,SpeI,PstI) which is used in Biobrick, we divided into plasmid of FliC and one of signal, and aimed the double transformation.

FliC-His generator

Experiment

• We regulate His-tagged FliC by lux promoter. Namely, if LuxR is expressed, bacteria can express FliC by Quorum Seinsing.
  
• Because the gene circuit of Quorum Sensing is on vector of colEI, we can use FliC and Quorum Sensing by using vector of p15A and do double transformation.

Results

Failed to ligate FliC-His generator and p15A origin vector.

• 混乱しています。ごめん。確認なんですが、FliC-His generatorをp15Aにのせるというのは、Biobrickにするためでないでしょうか。。僕の勘違いかな？。。そもそもQuorum sensing のベクターと、FliC-Hisチェックしてビーズ吸着させたときのベクターとを、ダブルトランスフォーメーションできないんだっけ？byとよたろ

FliC-his biobrick

Experiment

• Making Biobrick version of Flic-His and other stickly-FliCs. unfortunately, FliC has many EcoRI,SpeI,PstI sites. We have to eliminate them one by one by site-directed mutagenesis.

Results

Not yet (as of 10/26/2007). Cloning is still underway -->