Team:Todai-Tokyo/Notebook/bread

From 2009.igem.org

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Revision as of 00:57, 20 October 2009

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the notebook

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

10/1~13

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

10/15

ligate mtlD and plate1 7D

10/17

colony PCR of mtlD+plate1-7D

10/18

Miniprep of mtlD+plate1-7D
MtlD made a debut as an iGEM part!

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