Melbourne/Lab GV7
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| Vector | Insert | LIGATION | Transform | R= ESULT | PICKS | Plate | ||||||||||||||
| Bba_J61035 | digested | Antarctic phosphatas= e | heat inactivated |
gel purified | Source | dnpI digest |
gel purified | digested | Alkaline Phosphatase |
Antarctic phosphatas= e | heat inactivated |
gel purified | Type | vector qty | insert qty | amount | Target cells | #colonies day 1 |
||
| BU PCR | &X,P (1hr) | yes | extra digestion in gel (no good) | |||||||||||||||||
| P48J(1) | S,P | 80deg20m | yes | LL PCR | 1hr | yes | X,P 2hr | 80deg20m | yes | 5 min | 4uL | 4uL | all | XL-10 gold (45uL) | 10 (day2) | |||||
| P48J(1) | S,P | 80deg20m | yes | BU PCR | 1hr | 5 min | 4uL | 4uL | all | XL-10 gold (45uL) | 10 (day2) | BUA,BUB (21/9/07) | ||||||||
| P48J(1) | S,P | 80deg20m | yes | LL PCR | 1hr | yes | X,P 2hr | 80deg20m | yes | 5 min | 4uL | 25uL | all | XL-10 gold (45uL) | 10 (day2) | |||||
| P48J(1) | S,P | 80deg20m | yes | BU PCR | 1hr | 5 min | 4uL | 25uL | all | XL-10 gold (45uL) | 10 (day2) | BUC,BUD (21/9/07) | ||||||||
| P48J(1) | S,P | 80deg20m | yes | 4uL | all | XL-10 gold (45uL) | lots of colonies | |||||||||||||
| P48J(1) | S,P 1hr | 80deg20m | yes | LLPCR2 | 1h 30m | yes | X,P 2hr | 80deg20m | yes | 5 min | 2uL | 8uL(8ng/ul) | 20uL(22ng | Xl-10gold (20uL) | 100 | |||||
| P48J(1) | S,P 1hr | 80deg20m | yes | BUPCR2 | 1h 30m | yes | X,P 2hr | 80deg20m | yes | 5 min | 2uL | 8uL(8ng/ul) | 20uL(22ng | Xl-10gold (20uL) | 15 | |||||
| P48J(1) | S,P 1hr | 80deg20m | yes | 5min | 2uL | 20uL | Xl-10gold (20uL) | covered | ||||||||||||
| P48J(1) | S,P 1hr | 80deg20m | yes | LLPCR2 | 1h 30m | yes | X,P 2hr | 80deg20m | yes | 16hr 16deg | 2uL | 20uL | Xl-10gold (20uL) | 1000 | LLA,B,C,D,E | |||||
| P48J(1) | S,P 1hr | 80deg20m | yes | BUPCR2 | 1h 30m | yes | X,P 2hr | 80deg20m | yes | 16hr 16deg | 2uL | 20uL | Xl-10gold (20uL) | 300 | BUA,B,C,D,E 25/9/0 | digested in | ||||
| P48J(1) | S,P 1hr | 80deg20m | yes | 16hr 16deg | 2uL | 8uL(8ng/ul) | 20uL(22ng | Xl-10gold (20uL) | 300 | |||||||||||
| P48J(1) | S 1hr | complete digest | ||||||||||||||||||
| P48J(1) | P 1hr | incomplete digest | ||||||||||||||||||
| P48J(1) | P 1hr (in B3) | complete digest | ||||||||||||||||||
| P48J(1) | S,P overnight | yes | 2uL | 45 | ||||||||||||||||
| S,P overnight | yes | 5 min | 2uL | 23 | ||||||||||||||||
| S ovnight B2-> P (B3) 2hr | yes | 2uL | 48 | |||||||||||||||||
| S ovnight B2-> P (B3) 2hr | yes | 5 min | 2uL | 28 | ||||||||||||||||
| S only | yes | 5 min | 2uL | 3000 | ||||||||||||||||
| S only | yes | 2uL | 550 | |||||||||||||||||
| PUC18 ref | 5 min | 0.5uL | 84 | |||||||||||||||||
| PUC18 ref | ovnt | 0.5uL | 61 | |||||||||||||||||
| PUC18 ref | 0.5uL | 830 | ||||||||||||||||||
| P48J(1) | S ovnight B2-> P (B3) 2hr | yes | BUPCR4 | 1hr | yes | X,P 2h in (B2) then 2h in (B3) | 16hr 16de | 1uL | 12ul | 20ul | Xl-10gold (30uL) | fewer | 1:12A,B (5/10/07) | |||||||
| P48J(1) | S ovnight B2-> P (B3) 2hr | yes | BUPCR4 | 1hr | yes | X,P 2h in (B2) then 2h in (B3) | 16hr 16de | 2uL | 8ul | 20ul | Xl-10gold (30uL) | fewer | 2:8A,B | |||||||
| P48J(1) | S ovnight B2-> P (B3) 2hr | yes | BUPCR4 | 1hr | yes | X,P 2h in (B2) then 2h in (B3) | 16hr 16de | 2ul | 2ul | 20ul | Xl-10gold (30uL) | covered | 2:2A,B | |||||||
| P48J(1) | S ovnight B2-> P (B3) 2hr | yes | BUPCR4 | 1hr | yes | X,P 2h in (B2) then 2h in (B3) | 16hr 16de | 1ul | 4ul | 20ul | Xl-10gold (30uL) | covered | 1:4A,B | |||||||
| P48J(1) | S ovnight B2-> P (B3) 2hr | yes | BUPCR4 | 1hr | yes | X,P 2h in (B2) then 2h in (B3) | 16hr 16de | 2ul | 20ul | Xl-10gold (30uL) | covered | |||||||||
| P320I | X,P 5hrs | yes 1hr | 5min | 1ul | 6ul | 20ul | Xl-10gold (30uL) | fewer | ||||||||||||
| P320I | X,P 5hrs | yes 1hr | 5min | 2ul | 20ul | Xl-10gold (30uL) | covered | |||||||||||||
| P48J(2) | ES (5h40m) | BUPCR5 | 1 hr | yes | ES 2h | 2uL(11ng/ | 2ul(8ng/ul) | 2uL | Xl-10gold (45uL) | 400 | 60,61,62 | 1' | ||||||||
| P48J(2)(252ng/u= l | S (5h40m) | 15m @ 37deg <= /td> | 80deg 20min | BUPCR5 | 1 hr | yes | X,S 2h | 0.4uL(63n | 3uL(8ng/ul) | 2uL | Xl-10gold (45uL) | 96 | 63,64 | 2 | ||||||
| P320I(1)(154ng/= | ES (5h40m)<= /td> | BUPCR5 | 1 hr | yes | ES 2h | 2uL(9ng/ul) | 3ul(8ng/ul) | 2uL | Xl-10gold (45uL) | 37 | 3 | |||||||||
| P48J(2) | ES (5h40m)<= /td> | LLPCR3 | 1 hr | yes | ES 2h | 1uL (11ng/ | 4ul(5ng/ul) | 2uL | Xl-10gold (45uL) | 120 | 50,50,52 | 4 | ||||||||
| P48J(2) | S (5h40m) | 15m @ 37deg <= /td> | 80deg 20min | LLPCR3 | 1 hr | yes | X,S 2h | 0.3uL(63n | 4uL(5ng/ul) | 2uL | Xl-10gold (45uL) | 35 | 53,54 | 5 | ||||||
| P320I(1) | ES (5h40m)<= /td> | LLPCR3 | 1 hr | yes | ES 2h | 2uL(9ng/ul) | 4uL(5ng/ul) | 2uL | Xl-10gold (45uL) | 33 | 57,58,59 | 6 | ||||||||
| P48J(2) | ES (5h40m)<= /td> | 2uL (11ng/ul) | 2uL | Xl-10gold (45uL) | 60 | 71,72,73 | 7 | |||||||||||||
| P48J(2) | S (5h40m) | 15m @ 37deg <= /td> | 80deg 20min | 0.6uL(63ng/ul) | 2uL | Xl-10gold (45uL) | 35 | 8 | ||||||||||||
| P320I(1) | ES (5h40m)<= /td> | 3uL(9ng/ul) | 2uL | Xl-10gold (45uL) | 4 | 9 | ||||||||||||||
| P48J(2) | ES (5h40m) | BUPCR5 | 1 hr | yes | ES 2h | 2uL(11ng/ | 2ul(8ng/ul) | 2uL | DH5a(50uL) | 11 | 1' | |||||||||
| P48J(2)(252ng/u= l | S (5h40m) | 15m @ 37deg <= /td> | 80deg 20min | BUPCR5 | 1 hr | yes | X,S 2h | 0.4uL(63n | 3uL(8ng/ul) | 2uL | DH5a(50uL) | 18 | 65,66,67,68,69,70 | 2 | ||||||
| P320I(1)(154ng/= | ES (5h40m)<= /td> | BUPCR5 | 1 hr | yes | ES 2h | 2uL(9ng/ul) | 3ul(8ng/ul) | 2uL | DH5a(50uL) | 10 | 3 | |||||||||
| P48J(2) | ES (5h40m)<= /td> | LLPCR3 | 1 hr | yes | ES 2h | 1uL (11ng/ | 4ul(5ng/ul) | 2uL | DH5a(50uL) | 0 | 4 | |||||||||
| P48J(2) | S (5h40m) | 15m @ 37deg <= /td> | 80deg 20min | LLPCR3 | 1 hr | yes | X,S 2h | 0.3uL(63n | 4uL(5ng/ul) | 2uL | DH5a(50uL) | 5 | 55,56 | 5 | ||||||
| P320I(1) | ES (5h40m)<= /td> | LLPCR3 | 1 hr | yes | ES 2h | 2uL(9ng/ul) | 4uL(5ng/ul) | 2uL | DH5a(50uL) | 4 | 6 | |||||||||
| P48J(2) | ES (5h40m)<= /td> | 2uL (11ng/ul) | 2uL | DH5a(50uL) | 66 | 7 | ||||||||||||||
| P48J(2) | S (5h40m) | 15m @ 37deg <= /td> | 80deg 20min | 0.6uL(63ng/ul) | 2uL | DH5a(50uL) | 0 | 8 | ||||||||||||
| P320I(1) | ES (5h40m)<= /td> | 3uL(9ng/ul) | 2uL | DH5a(50uL) | 11 | 9 |
