Team:Macquarie Australia/Protocols/GibsonAssembly

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Revision as of 10:32, 5 September 2012

Gibson Assembly Protocol

Before entering the lab, the assigned Gibson Assembly team for the day organised agar plate requirements, concentration calculations and equipment requirements.

Each tube was labelled with antibiotic (C = chloramphenicol, K = kanamycin, A = ampicillin).

Instruction	#1C style="background-color: yellow;"\|	#2K	#2A	#3K	#3A	#4C	#5K	#5A
1. Addition of 20ng Gene Block Fragments in appropriate tube	2µl Hemo_T7A	2µl Hemo_A	2µl Hemo_A	2µl Deino_A	2µl Deino_A	2µl Agro_T7A	2µl Agro_A	2µl Agro_A
	2µl Hemo_B	2µl Hemo_B	2µl Hemo_B	2µl Deino_B	2µl Deino_B	2µl Agro_B	2µl Agro_B	2µl Agro_B
	nil	nil	nil	2µl Deino_C	2µl Deino_C	2µl Agro_C	2µl Agro_C	2µl Agro_C
	nil	nil	nil	2µl Deino_D	2µl Deino_D	2µl Agro_D	2µl Agro_D	2µl Agro_D
	nil	nil	nil	2µl Deino_E	2µl Deino_E	2µl Agro_E	2µl Agro_E	2µl Agro_E
2. Addition of 0.05 pmol of vector	2.7µl PSB-1C3	2.9µl PSB-1K3	2.8µl PSB-1A3	2.9 µl PSB-1K3	2.8 µl PSB-1A3	2.7µl PSB-1C3	2.9µl PSB-1K3	2.8µl PSB-1A3
3. Addition of Gibson Master Mix (µl)	10	10	10	12.9	12.8	12.7	12.9	12.8
4. Addition of deionised H2O	3.3µl	3.1µl	3.2µl	nil	nil	nil	nil	nil

@@ Line 11: / Line 11: @@
 |-
 ! scope="col" colspan="1" | Instruction
-! scope="col"| #1C
+! scope="col"| #1C style="background-color: yellow;"|
 ! scope="col"| #2K
 ! scope="col"| #2A