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        <h2 style="color:black" align="center"><b>Background</b></h2>
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        <p>With the expanding of human settlements and the development of civil engineering, the demand of novel cementation material is increasing rapidly. To echo such demand, we developed Euk.cement: a live eukaryotic cell based auto-cementation kit. By surface displayed silica binding peptides and secreted flocculating proteins, Euk.cement will target onto any silica containing particles, such as sands and rocks, and stick them together. This system will be automatically initiated only in dark with a light operated switch. While carbon dioxide released from the metabolism of cells will finally complete the calcium carbonate sedimentation. This economical and ecological friendly innovation can be utilized for a wide range of industrial or environmental applications, such as construction and restoration of building foundations, bridge piers, or even artificial reefs for aquaculture. Since we have exploited a new kind of marine yeast as chassis, the application field of eukaryotic synthetic biology has then been considerably broadened.  
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    <p>To echo such demand, we developed Euk.cement: a live eukaryotic cell based auto-cementation kit. Euk.cement is supposed to target onto any silica containing particles, such as sands and rocks, and stick them together. And conditions permitting,with the addition of Ca2+ in our substrate,carbon dioxide released from the metabolism of cells will together making calcium carbonate sedimentation to final enhance the consolidation progress.
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          Euk.cement owns an abroad industrial or environmental application’s prospect, such as construction and restoration of building foundations, bridge piers, or even artificial reefs for aquaculture.
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Revision as of 12:13, 17 September 2015

<!DOCTYPE html> description


Euk.cement
A live eukaryotic cell based auto-cementation kit



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Details

Standalization

Week 1 6.29-7.5

Week 2 7.6-7.12

Week 3 7.13-7.19

Week 4 7.20-7.26

Week 5 7.27-8.2

Week 6 8.3-8.9

PCR: More details, protocol WangJie Liu and JunJie Xu

Restriction Enzyme Digestion
WangJie Liu and JunJie Xu

DNA ligation&Transformation
WangJie Liu and JunJie Xu

Successfully align the sequencing results with the objective sequences:
Colony PCR
WangJie Liu and JunJie Xu

Extract the plasmid,Conserving.

Week 7 8.10-8.16

PCR:
More details, protocol
WangJie Liu

Restriction Enzyme Digestion
WangJie Liu

DNA ligation&Transformation
WangJie Liu

Colony PCR
Successfully align the sequencing results with the objective sequences:
WangJie Liu

Extract the plasmid,Conserving.

Week 8 8.17-8.23

Week 9 8.24-8.30

Week 10 8.31-9.6

Successfully align the sequencing results with the objective sequences
Standalization of BD-CRY2 and AD-CIB1 finished.

Successfully align the sequencing results with the objective sequences
Standalization of CRY2 and CIB1 finished.

Light Control

Week 1 6.29-7.5

Week 2 7.6-7.12

Successfully align the sequencing results with the objective sequences of Pgal1 and CYC_TT.
Extract the plasmid.
Conserving.

Week 3 7.13-7.19

Week 4 7.20-7.26

Week 5 7.27-8.2

Successfully align the sequencing results with the objective sequences

Digestion Test
Guozhao Wu

Week 6 8.3-8.9

Successfully align the sequencing results with the objective sequences
Pgal-ROX1-CYC_TT circuit was constructed.

Week 7 8.10-8.16

Week 8 8.17-8.23

Week 9 8.24-8.30

Week 10 8.31-9.6

viscous parts

Week 1 6.29-7.5

Week 2 7.6-7.12

PCR:
More details, protocol
Wang Xi and JunJie Xu

Restriction Enzyme Digestion
Wang Xi and JunJie Xu

DNA ligation&Transformation
Wang Xi and JunJie Xu

Colony PCR
Successfully align the sequencing results with the objective sequences:
Wang Xi and JunJie Xu

Extract the plasmid,Conserving

Week 3 7.13-7.19

PCR:
More details, protocol
ZhangYu Cheng

Restriction Enzyme Digestion
ZhangYu Cheng

DNA ligation&Transformation
ZhangYu Cheng

Colony PCR
ZhangYu Cheng
Successfully align the sequencing results with the objective sequences:

Extract the plasmid,Conserving.

Week 4 7.20-7.26

PCR:
More details, protocol
Wang Xi and JunJie Xu

Restriction Enzyme Digestion
Wang Xi and JunJie Xu

DNA ligation&Transformation
Wang Xi and ZhangYu Cheng

Colony PCR
Wang Xi and JunJie Xu
Successfully align the sequencing results with the objective sequences:

Extract the plasmid,Conserving.

Tramsformation to yeast
Lei Yin

Genome extract and PCR test
Lei Yin

Positive strain conservation

Week 5 7.27-8.2

PCR:
More details, protocol
ZhangYu Cheng

Restriction Enzyme Digestion
ZhangYu Cheng

DNA ligation&Transformation
ZhangYu Cheng

Colony PCR
ZhangYu Cheng
Successfully align the sequencing results with the objective sequences:

Extract the plasmid,Conserving.

Trasnsform to yeast

Genome extract and PCR test
Lei Yin

Positive strain conservation

Week 6 8.3-8.9

PCR:
More details, protocol
ZhangYu Cheng

Restriction Enzyme Digestion
ZhangYu Cheng

DNA ligation&Transformation
ZhangYu Cheng

Colony PCR
ZhangYu Cheng
Successfully align the sequencing results with the objective sequences:

Extract the plasmid,Conserving.

Trasnsform to yeast
Lei Yin

Genome extract and PCR test
Lei Yin

Positive strain conservation

Week 7 8.10-8.16

SDS-PAGE
Lei Yin

Immunoflourecense
ZhangYu Cheng

Verification of the secreting Mcfp-3 (Coomassie blue staining)
ZhangYu Cheng

Week 8 8.17-8.23

Week 9 8.24-8.30

Week 10 8.31-9.6

Improvement

Week 1 6.29-7.5

Week 2 7.6-7.12

Week 3 7.13-7.19

Week 4 7.20-7.26

Week 5 7.27-8.2

Week 6 8.3-8.9

Week 7 8.10-8.16

Week 8 8.17-8.23

Week 9 8.24-8.30

Week 10 8.31-9.6

Consolidation

Week 1 6.29-7.5

Week 2 7.6-7.12

Week 3 7.13-7.19

Week 4 7.20-7.26

Week 5 7.27-8.2

Week 6 8.3-8.9

Week 7 8.10-8.16

Week 8 8.17-8.23

Week 9 8.24-8.30

Week 10 8.31-9.6

Host strain

We adopt a marine yeast Yarrowia lipolytca JMY1212 as our chassis for the specific application this year--building artificial reef. Our host is generally regarded as safe microorganism and has an abroad application prospect in industries of foods and medicine. Researches shows that Y.lipolytica is widely found in marine environment which exists stably in the ocean and healthy marine fishes’ intestine and it is able to make use of Glucose,ethanol,acetate or other hydrophobic substrate(alkanes,fatty acid and lipid ) as carbon source with fast growing capacity and adaptability for high-density fermentation. Moreover in the expression system of Y.lipolytica, transformed plasmids can integrate into multiple site and they are all genetically stable in the genome. And the signal peptide XPR2 pre,LIP2 prepro and so on are all demonstrated to own high efficiency of secreting protein.So it is obvious Y.lipolytica is a powerful host to express and secrete complex recombinant protein. Based on the above reasons,Yarrowia lipolytica are regarded as ideal strain for our marine application of micro-solidation

Improvement

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