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Team:Todai-Tokyo/Notebook/bread
From 2009.igem.org
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1.Clone the Glu1 (glucoamylase) from Saccharomycopsis fibuligera and insert it in the yeast chromosome by homologous recombination<BR> | 1.Clone the Glu1 (glucoamylase) from Saccharomycopsis fibuligera and insert it in the yeast chromosome by homologous recombination<BR> | ||
2-a Clone mtlD (mannitol synthase) from E.coli and insert it in the yeast chromosome by homologous recombination<BR> | 2-a Clone mtlD (mannitol synthase) from E.coli and insert it in the yeast chromosome by homologous recombination<BR> | ||
| - | Replace gpd1 gene by mtlD and gpd2 gene by glu1<BR> | + | Replace gpd1 gene by mtlD and gpd2 gene by glu1<BR> |
2-b Clone xylose isomerase gene and D-tagatose 3-epimerase gene from Mesorhizobium loti.<BR> | 2-b Clone xylose isomerase gene and D-tagatose 3-epimerase gene from Mesorhizobium loti.<BR> | ||
| - | Replace gpd1 gene by D-tagatose 3-epimerase gene and gpd2 gene by glu1. Transform a plasmid coding xylose isomerase gene. | + | Replace gpd1 gene by D-tagatose 3-epimerase gene and gpd2 gene by glu1. Transform a plasmid coding xylose isomerase gene. |
=='''~9/20'''== | =='''~9/20'''== | ||
Revision as of 03:51, 16 October 2009
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Contents |
Plan
Aim:Create yeast that can be used to make sweet and low energy bread
Methods:
1.Clone the Glu1 (glucoamylase) from Saccharomycopsis fibuligera and insert it in the yeast chromosome by homologous recombination
2-a Clone mtlD (mannitol synthase) from E.coli and insert it in the yeast chromosome by homologous recombination
Replace gpd1 gene by mtlD and gpd2 gene by glu1
2-b Clone xylose isomerase gene and D-tagatose 3-epimerase gene from Mesorhizobium loti.
Replace gpd1 gene by D-tagatose 3-epimerase gene and gpd2 gene by glu1. Transform a plasmid coding xylose isomerase gene.
~9/20
- PCR of mtlD
- PCR of gpd1 promoter
- TA cloning of mtlD
9/21
- Miniprep of mtlD
9/22
- read the sequence of mtlD→successful
- mtlD primers with HAtag come
9/25
- PCR of mtlD with new primer→failed
9/26
- PCR of mtlD with new primer→successful
9/27
- PCR of gpd1 promoter with Pfu Ultra
October
- PCR of Glu1
- TA cloning of Glu1
- PCR of gpd1 promoter with ExTaq
10/14
- cut mtlD and plate1 7D both by EcoRI and PstI
- colony PCR of Glu1
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