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From 2010.igem.org

Contents 1 Lab work 1.1 Alkane degration 1.2 Hydrocarbon Sensing 1.2.1 Ligation

Lab work

Alkane degration

Yesterday's plates showed colonies. To check whether these contained the desired insert a colony PCR was performed. The products of this PCR were checked on a gel.

On the gel, two colonies of 011A showed the right length. None of the lengths for 013A or K were correct. They did however match the PCR length (009A, 482) of one the inserts which was digested in order to create this BioBrick. Next week we will take a few more colonies from the plate and check these, and otherwise the digestion and ligation will be tried again.

Lane description

#	Description	Primers	Expected length (bp)	✓	✗
L	SmartLadder	n/a	n/a	n/a	n/a
1-5	011A	G00100 + G00101	1816	1,5	2,3,4
6-10	013A	G00100 + G00101	1929	none	6,7,8,9,10
11-15	013K	G00100 + G00101	1929	none	11,12,13,14,15
L	SmartLadder	n/a	n/a	n/a	n/a

Hydrocarbon Sensing

Ligation

Digested pAlkS, B0032, AlkS and P(Caif) were ligated as followed:

#	Description	Volume (µl)
1	PalkS12-B0032	9.5
	AlkS-E0240	3.0
	ligase	1.5
	ligase buffer	1
2	PalkS12-B0032	10.5
	pSB1T3	2.0
	ligase	1.5
	ligase buffer	1
3	P(Caif)-B0032	9.5
	AlkS-E0240	3.0
	ligase	1.5
	ligase buffer	1
4	P(Caif)-B0032	10.5
	pSB1T3	3.0
	ligase	1.5
	ligase buffer	1

The ligation products were checked on a gel.