plasmidRepairReactions

	A	B	C	D	E	F	G	H	I
1	Plasmid	Concentration	Desired nicked mass (ug)	Desired ligated mass (ug)	Desired Supercoiled)	gel volume
2	pFC9-T7-init-VR14-2-midi-D	228.817484	3	2	1	0.437029541	4.562970459
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4
5
6	NtBspQI Rxn
7	npH20	r3.1 10x Buffer	Plasmid	Plasmid volume (ul)	Nt.BspQI (ul)	Reaction volume (ul)	concentration	Gel Volume
8	8.389113771	2.5	pFC9-T7-init-VR14-2-midi-D	13.11088623	1	25	120	0.8333333333	4.166666667
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11	Taq Ligase Rxn
12	Volume nicked substrate	npH20	Taq Ligase 10x Buffer	Total volume (ul)	Taq ligase	Template concentration	Gel volume
13	16.667	26.333	5	50	2	40.0008	2.499950001	2.500049999
14
15	Gyrase Rxn
16	Volume ligated substrate	npH20	5x Topogen Gyrase buffer	Gyrase (5u/ul) ul		Total Volume	Concentration	Aliquote mass
17	24.99950001	13.00049999	10	2		50	20	100
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19
20	Gyrase Rxn time points	Aliquote volume (ul)
21	10	5
22	15	5
23	20	5	Total removed	Total remaining
24	30	5	40	10
25	40	5
26	50	5
27	60	5
28	80	5
29	120	5
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33	Gel (0.8% 1x TAE SeaPlaque agarose)
34	Lane	Sample	Gyrase Exposure time	Voltage	Mass (ng)
35	1	pFC9VR14	0	60	100
36	2	PFC9VR14 + NtBspQI	0	60	100
37	3	PFC9VR14 + NtBspQI + Taq ligase	0	60	100
38	4	PFC9VR14 + NtBspQI + Taq ligase	10	60	100
39	5	PFC9VR14 + NtBspQI + Taq ligase	15	60	100
40	6	PFC9VR14 + NtBspQI + Taq ligase	20	60	100
41	7	PFC9VR14 + NtBspQI + Taq ligase	30	60	100
42	8	PFC9VR14 + NtBspQI + Taq ligase	40	60	100
43	9	PFC9VR14 + NtBspQI + Taq ligase	50	60	100
44	10	PFC9VR14 + NtBspQI + Taq ligase	60	60	100
45	11	PFC9VR14 + NtBspQI + Taq ligase	80	60	100
46	12	PFC9VR14 + NtBspQI + Taq ligase	120	60	100
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