Part:BBa_K539691:Experience

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Experience by 2011 iGEM NCTU_FORMOSA

Figure.1 Firstly , we ligase B0030 with ilvC.

Figure.2 At the sametime , we ligase B0030 with ilvD.

Figure.3 Meanwhile , we ligase B0032 with Alss.

Figure.4 To make sure we obtain the correct part , colony PCR is needed.

Figure.5 Secondly , we ligase Plac with B0032+Alss

Figure.6 Thirdly , we ligase Plac+B0032+Alss with B0030+ilvC

Figure.7 At the same time , we ligase B0030+ilvD with RNA thermometer+tetR+double terminator

Figure.8 Finally , we ligase the whole circuit:promoter(lacI regulated)+Alss+ilvC+ilvD(each preceded by own RBS)and RNA thermometer+terminator

We cotransformed Part:BBa_K539691 and Part:BBa_K539742 into DH5α, so the expression of Alss, ilvC, ilvD and kivd are under the control of temperature.

And to realize whether Alss,ilvC and ilvD have any impact to the production of isobutanol, we compare the circuit which contain Alss,ilvC and ilvD genes with the one with only a single kivd gene in it.

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Figure.9 Additionally insert the following genes, alss, ilvC, and ilvD(three precursory genes), the production of isobutanol will extremely increase. (About 80 times)

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