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From 2014.igem.org

Contents 1 LabWork 1.1 B- Construction of the fusion protein 1.2 D- Lemon scent 1.2.1 PCR LS 1.2.2 Ligation 1.2.3 pPS5 1.3 pPS3 and pPS4

LabWork

B- Construction of the fusion protein

We made a classic exttraction of plasmids from the liquid cultures.

Check by electrophoresis if it worked and send it for sequencing.

D- Lemon scent

by Mélanie

PCR LS

Electrophoresis of the PCR made Yesterday

[photo]

The PCR have success so I use the PCR clean up kit to purify it

[photo] Electrophoresis after purification

Ligation

We already have some pPSI digested and dephosphorelated

So I do a ligation:

component	volume
H2O	5μl
buffer	2μl
ligase	1μl
pPSI	2μl
LS PCR	10μl

2 hours at room temperature and over night at 4°

pPS5

Plasmid exctraction using the kit digestion by SalI

component	volume
H2O	11μl
buffer	5μl
SalI	2μl
pPS5	30μl

Electrophoresis

[photo]

We saw that we have something strang with our plasmid

pPS3 and pPS4

We digest the plasmid by HindIII to direct our insert

! scope=col | component ! scope=col | volume |- |H₂O |6μl |- |buffer |1μl |- |HindIII |1μl |- |pPS3/4 |2μl |}

[photo]

results are very strange