Team:Tokyo Metropolitan/Notebook/S16

From 2011.igem.org

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(Created page with "{{:Team:Tokyo_Metropolitan/Header}} <html> <div style="width: 800px; margin-left: 70px; padding-top: 25px; padding-left: 20px;"> </html> = -Visualization- = @plasmid extraction...")
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@electrophoresis for confirmation, but could not find bind.
@electrophoresis for confirmation, but could not find bind.
 +
 +
 +
= -Sequencing- =
 +
 +
 +
@Killer＋Target
 +
 +
①J23100-K317037-K283030　/　VF
 +
 +
could read 500 base which contain vector and the first half of J23100 and K317037, but not bond to ②
 +
 +
 +
②J23100-K317037-K283030　/　VR
 +
 +
could read 500 base which contain vector and the latter half of K283030,
 +
 +
but could not bond to ①, deficiene 50 base from terminator codon of CheZ in K283030 (also terminator codon).
 +
 +
 +
 +
③P0140　/　VF
 +
 +
④P0140　/　VR
 +
 +
could not find both of the sequence
 +
 +
 +
@Visual
 +
 +
⑤OriT_GFP_OriT_1　/　VF
 +
 +
could read about 350 base and confirmate vector and part of OriT for 60 base
 +
 +
 +
⑥OriT_GFP_OriT_1　/　GFP
 +
 +
⑦OriT_GFP_OriT_1　/　VR
 +
 +
could read about 650 base from ⑥ and about 350 base from ⑦,
 +
 +
and bond each sequence to one
 +
 +
 +
 +
⑧OriT_GFP_OriT_2　/　VF
 +
 +
could read about 350 base and confirmate vector and part of OriT for 60 base
 +
 +
 +
⑨OriT_GFP_OriT_2　/　GFP
 +
 +
could read about 600 base, but that is just sequence of GFP
 +
 +
 +
⑩OriT_GFP_OriT_2　/　VR
 +
 +
could read 300 base, but not agree with OriT_GFP_OriT
 +
 +
 +
⑪OriT_GFP_OriT_3　/　VF
 +
 +
could read 250 base which contain OriT for 60 base and vector for other sequence
 +
 +
 +
⑫OriT_GFP_OriT_3　/　GFP
 +
 +
could not read sequence
 +
 +
 +
⑬OriT_GFP_OriT_3　/　VR
 +
 +
could read about 300 base and find sequence of OriT
 +
 +
 +
⑭OriT_GFP_OriT_4　/　VF
 +
 +
could read 200 base and confirmated vector and the first half of OriT
 +
 +
 +
⑮OriT_GFP_OriT_4　/　GFP
 +
 +
could read about 600 base, but that is just sequence of GFP
 +
 +
 +
⑯OriT_GFP_OriT_4　/　VR
 +
 +
could read about 300 base and confirmated the latter half of OriT
 +
 +
 +
@summary
 +
 +
Targeting
 +
 +
succeed in ligation, but deficiene 50 base from terminator codon of CheZ in K28303.
 +
 +
I guessed there is not His-tag.
 +
 +
could not read sequence of P0140.
 +
 +
decrease tenpate concentration and sequencing again
 +
 +
 +
Visual
 +
 +
4 sumples of OriT-GFP-OriT was turned out wrong.
 +
 +
1 and 4 were OriT-GFP.
 +
 +
2 was unknown.
 +
 +
3 was OriT only, which is self ligation.

Revision as of 11:37, 5 October 2011

-Visualization-

@plasmid extraction oriTr-GFP-oriTr and JM109

@plasmid extraction and DNA refine oriTr-GFP-oriTr and JM109

@DNA densitometry

oriTr-GFP-oriTr 5 26.1 ng/μl

oriTr-GFP-oriTr 6 51.3 ng/μl

oriTr-GFP-oriTr 7 47.2 ng/μl

oriTr-GFP-oriTr 8 58.4 ng/μl

@restriction enzyme to oriTr-GFP-oriTr (XbaⅠ, PstⅠ)

@electrophoresis and gel extraction oriTr-GFP-oriTr5～8 and pSB1C3(restriction enzymed 9/15)

@DNA densitometry

totaled oriTr-GFP-oriTr5,6,8 18.4 ng/μl

pSB1C3① 16.5 ng/μl

pSB1C3② 11.5 ng/μl

@electrophoresis for confirmation, but could not find bind.


-Sequencing-

@Killer＋Target

①J23100-K317037-K283030　/　VF

could read 500 base which contain vector and the first half of J23100 and K317037, but not bond to ②


②J23100-K317037-K283030　/　VR

could read 500 base which contain vector and the latter half of K283030,

but could not bond to ①, deficiene 50 base from terminator codon of CheZ in K283030 (also terminator codon).


③P0140　/　VF

④P0140　/　VR

could not find both of the sequence


@Visual

⑤OriT_GFP_OriT_1　/　VF

could read about 350 base and confirmate vector and part of OriT for 60 base


⑥OriT_GFP_OriT_1　/　GFP

⑦OriT_GFP_OriT_1　/　VR

could read about 650 base from ⑥ and about 350 base from ⑦,

and bond each sequence to one


⑧OriT_GFP_OriT_2　/　VF

could read about 350 base and confirmate vector and part of OriT for 60 base


⑨OriT_GFP_OriT_2　/　GFP

could read about 600 base, but that is just sequence of GFP


⑩OriT_GFP_OriT_2　/　VR

could read 300 base, but not agree with OriT_GFP_OriT


⑪OriT_GFP_OriT_3　/　VF

could read 250 base which contain OriT for 60 base and vector for other sequence


⑫OriT_GFP_OriT_3　/　GFP

could not read sequence


⑬OriT_GFP_OriT_3　/　VR

could read about 300 base and find sequence of OriT


⑭OriT_GFP_OriT_4　/　VF

could read 200 base and confirmated vector and the first half of OriT


⑮OriT_GFP_OriT_4　/　GFP

could read about 600 base, but that is just sequence of GFP


⑯OriT_GFP_OriT_4　/　VR

could read about 300 base and confirmated the latter half of OriT


@summary 

Targeting

succeed in ligation, but deficiene 50 base from terminator codon of CheZ in K28303.

I guessed there is not His-tag.

could not read sequence of P0140.

decrease tenpate concentration and sequencing again 


Visual

4 sumples of OriT-GFP-OriT was turned out wrong.

1 and 4 were OriT-GFP.

2 was unknown.

3 was OriT only, which is self ligation.

Retrieved from "http://2011.igem.org/Team:Tokyo_Metropolitan/Notebook/S16"