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Edinburgh/Notebook/PCR products
From 2008.igem.org
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'''P9:''' ''crtI'' (from Douglas's maxiprep of the mutated ''crtIB'' plasmid). (08.07.08)<br /> | '''P9:''' ''crtI'' (from Douglas's maxiprep of the mutated ''crtIB'' plasmid). (08.07.08)<br /> | ||
'''P10:''' ''crtY'' (from ''P. ananatis'' cells). Failed. (08.07.08)<br /> | '''P10:''' ''crtY'' (from ''P. ananatis'' cells). Failed. (08.07.08)<br /> | ||
| - | '''P11:''' fusion PCR | + | '''P11:''' fusion PCR of rbs+''dxs''. Failed. (10.07.08)<br /> |
| - | '''P12:''' repeat fusion PCR | + | '''P12:''' repeat fusion PCR of rbs+''dxs'', using L11 and primers rbs2fclon1+pSB1A2insr1. (13.07.08)<br /> |
'''P13 and P14:''' MABEL mutation of site 1 on ''glgC''-mut2 clones M19 and M22. (14.07.08)<br /> | '''P13 and P14:''' MABEL mutation of site 1 on ''glgC''-mut2 clones M19 and M22. (14.07.08)<br /> | ||
'''P15:''' fusion PCR for rbs+''appY'' using L15 as a template. (15.07.08)<br /> | '''P15:''' fusion PCR for rbs+''appY'' using L15 as a template. (15.07.08)<br /> | ||
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'''P20 to P23:''' second attempts at ''cenA'', ''cenB'', ''cenC'' and ''cex''. (18.07.08) - Failed<br /> | '''P20 to P23:''' second attempts at ''cenA'', ''cenB'', ''cenC'' and ''cex''. (18.07.08) - Failed<br /> | ||
'''P24, P25:''' repeat PCR for ''cenA'' and ''cex'' using Pfu. (19.07.08) - Failed<br /> | '''P24, P25:''' repeat PCR for ''cenA'' and ''cex'' using Pfu. (19.07.08) - Failed<br /> | ||
| - | '''P26:''' Positive control PCR with primers fd1 and rd1 to amplify 16S rRNA gene ''rrnB'' from '' | + | '''P26:''' Positive control PCR with primers fd1 and rd1 to amplify 16S rRNA gene ''rrnB'' from ''C. fimi''. (22.07.08: AM) - Failed<br /> |
| - | '''P27:''' ''crtY'' from ''P. ananatis'' cells. - Failed<br /> | + | '''P27:''' ''crtY'' from ''P. ananatis'' cells. (23.07.08: AM, Yan, OG) - Failed<br /> |
| - | '''P28 | + | '''P28:''' fusion PCR of rbs+''crtB'' from L18. (23.07.08: AM, Yan, OG) |
| - | '''P30:''' | + | '''P29:''' fusion PCR of rbs+''crtI'' from L19. (23.07.08: AM, Yan, OG)<br /> |
| + | '''P30:''' repeat positive control of ''rrnB'' from ''C. fimi'', annealing 1 minute. (23.07.08: AM)<br /> | ||
'''P31:''' pZntA promoter<br /> | '''P31:''' pZntA promoter<br /> | ||
| - | '''P32-P35:''' repeat PCR | + | '''P32-P35:''' repeat PCR of ''cenA'', ''cenB'', ''cenC'' and ''cex'', annealing 1 minute. (24.07.08: AM) - Failed<br /> |
'''P36:''' M43 (''glgC''-mut1,2) with primers glgCf2/glgCr2 (25.07.08: AM)<br /> | '''P36:''' M43 (''glgC''-mut1,2) with primers glgCf2/glgCr2 (25.07.08: AM)<br /> | ||
'''P37:''' ''crtY'' from ''P. ananatis''. (25.07.08: AM) - Failed<br /> | '''P37:''' ''crtY'' from ''P. ananatis''. (25.07.08: AM) - Failed<br /> | ||
Revision as of 11:57, 26 August 2008
P1: dxs from Escherichia coli JM109. (25.06.08)
P2: appY from E. coli JM109. (25.06.08)
P3: glgC from E. coli JM109. (25.06.08)
P4: fusion PCR for BABEL1+glgC. (01.07.08)
P5: fusion PCR for BABEL2+glgC. (01.07.08)
P6: mutagenic PCR to remove EcoRI site 1 from glgC (M11). (04.07.08)
P7: mutagenic PCR to remove EcoRI site 2 from glgC (M11). (04.07.08)
P8: crtB (from Pantoea ananatis cells). (08.07.08)
P9: crtI (from Douglas's maxiprep of the mutated crtIB plasmid). (08.07.08)
P10: crtY (from P. ananatis cells). Failed. (08.07.08)
P11: fusion PCR of rbs+dxs. Failed. (10.07.08)
P12: repeat fusion PCR of rbs+dxs, using L11 and primers rbs2fclon1+pSB1A2insr1. (13.07.08)
P13 and P14: MABEL mutation of site 1 on glgC-mut2 clones M19 and M22. (14.07.08)
P15: fusion PCR for rbs+appY using L15 as a template. (15.07.08)
P16 to P19: first attempts at cenA, cenB, cenC and cex. (17.07.08) - Failed
P20 to P23: second attempts at cenA, cenB, cenC and cex. (18.07.08) - Failed
P24, P25: repeat PCR for cenA and cex using Pfu. (19.07.08) - Failed
P26: Positive control PCR with primers fd1 and rd1 to amplify 16S rRNA gene rrnB from C. fimi. (22.07.08: AM) - Failed
P27: crtY from P. ananatis cells. (23.07.08: AM, Yan, OG) - Failed
P28: fusion PCR of rbs+crtB from L18. (23.07.08: AM, Yan, OG)
P29: fusion PCR of rbs+crtI from L19. (23.07.08: AM, Yan, OG)
P30: repeat positive control of rrnB from C. fimi, annealing 1 minute. (23.07.08: AM)
P31: pZntA promoter
P32-P35: repeat PCR of cenA, cenB, cenC and cex, annealing 1 minute. (24.07.08: AM) - Failed
P36: M43 (glgC-mut1,2) with primers glgCf2/glgCr2 (25.07.08: AM)
P37: crtY from P. ananatis. (25.07.08: AM) - Failed
P38~P41: cenA, cenB, cenC and cex from heat-killed cell suspension.
P42~P45: cenA, cenB, cenC and cex from 'impure' DNA solution.
P46~P47: cenA and cex from 'impure' DNA solution, annealing 65C. Failed.
P48: crtY (new primer mixture) (previously P38a) (30.07.08: CF) - Failed
P49: Recreation of P12 (rbs+dxs) (previously P39a) (30.07.08: CF)
P50: third mutagenesis of glgc-mut1,2 (04.08.08: YAN)
P51: pCstA from E. coli cells (06.08.08: CF) - Failed
P52: cenA from heat killed C. fimi (07/08/08: YAN, AM) - Failed
P53: cex from heat killed C. fimi (07/08/08: YAN, AM) - Failed
P54: pCstA from E. coli cells - Purified (07.08.08: CF)
P55: PCR products of CenA by KOD
P56: PCR products of CenA by velocity
P57: PCR products of CeX by KOD
P58: PCR products of CeX by Velocity
P59: pSB1A2-crtY (09.08.08: CF)
P60: pSB1A2-rbs+crtY (09.08.08: CF)
P61: 16s rRNA rrnB from C. fimi genomic DNA - successful(10.08.08: CF)
P62: cenA from C. fimi genomic DNA - failed (10.08.08: CF)
P63: cex from C. fimi genomic DNA - failed (10.08.08: CF)
P64: cex from C. fimi genomic DNA, w/ glycerol, annealing 60°C. Success (11.08.08: AM)
P65: cenA from C. fimi genomic DNA, w/ glycerol, annealing 56°C/20s. Success (12.08.08: AM)
P66: zm1 (ISA1) (12.08.08: CF)
P67: zm2 (ISA2) (12.08.08: CF)
P68: cenB from C. fimi genomic DNA. Failed (13.08.08: AM)
P69: cenC from C. fimi genomic DNA. Failed (13.08.08: AM)
P70: M43 (glgC mut 1+2). Failed (14.08.08: AM)
P71: M120 (glgC mut 1+2+3). Successful (14.08.08: AM)
P71A: Repeat M43 (glgc mut 1+2). Succeessful (15.08.08: AM)
P72: Repeat M120 (glgC mut 1+2+3). Successful (15.08.08: AM)
P73~P77: rrnB from C. fimi genomic DNA purified on 14.08.08. P74 possibly successful (15.08.08: AM)
P78A: rrnB from original C. fimi genomic DNA as positive control. Failed (15.08.08: AM)
P78: SOB2-glgC mut 1+2 from L45 (19.08.08: AM)
P79: SOB2-glgC mut 1+2+3 from L46 (19.08.08: AM)
P80: SOB2-glgC mut 1+2+3 from L47 (19.08.08: AM)
P81: SOB2-rbs-glgc mut 1+2 from L45 (19.08.08: AM)
P82: SOB2-rbs-glgC mut 1+2+3 from L46 (19.08.08: AM)
P83: SOB2-rbs-glgC mut 1+2+3 from L47 (19.08.08: AM)
P84: zm1 (ISA1) from X11 (20.08.08: AM)
P85: zm2 (ISA2) from X12 (20.08.08: AM)
