SBB09Ntbk-Madhvi Venkatesh: Difference between revisions
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<pre> | <pre> | ||
Construction of {<fliD>} (C-terminal) | Construction of {<fliD>} (C-terminal type 1) | ||
PCR Omv044F/Omv045R on MG1655 genomic DNA (485bp, gp=A) | PCR Omv044F/Omv045R on MG1655 genomic DNA (485bp, gp=A) | ||
PCR Omv045F/Omv046R on MG1655 genomic DNA (970bp, gp=B) | PCR Omv045F/Omv046R on MG1655 genomic DNA (970bp, gp=B) | ||
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---- | ---- | ||
Omv044F Forward Bglbricking of {<fliD!} ccagtGAATTCatgAGATCTgcaagtatttcatcgctggg | Omv044F Forward Bglbricking of {<fliD!} ccagtGAATTCatgAGATCTgcaagtatttcatcgctggg | ||
Omv046R Reverse Bglbricking of {<fliD>} gcagtGGATCCcttggaattactgttgttttcg | |||
Omv045F Forward Removing BamHI site in {<fliD!} cgttaagcggCatccgtgatgcc | |||
Omv045R Reverse Removing BamHI site in {<fliD!} ggcatcacggatGccgcttaacg | |||
</pre> | |||
<pre> | |||
Construction of {a~fliD>} (C-terminal type 2) | |||
PCR Omv044F/Omv045R on MG1655 genomic DNA (492bp, gp=A) | |||
PCR Omv045F/Omv046R on MG1655 genomic DNA (970bp, gp=B) | |||
---- | |||
PCR Omv044F/Omv046R on A + B (1439bp, EcoRI/BamHI) | |||
Sub into pBca9495CA-Bca1144#5 (EcoRI/BamHI, 3039+910, L) | |||
Product is pBca9495KC-M10001 {a~fliD>} | |||
---- | |||
Omv047F Forward Bglbricking of {a~fliD!} ccagtGAATTCatgAGATCTtcgcatggcaagtatttcatcgctggg | |||
Omv046R Reverse Bglbricking of {<fliD>} gcagtGGATCCcttggaattactgttgttttcg | Omv046R Reverse Bglbricking of {<fliD>} gcagtGGATCCcttggaattactgttgttttcg | ||
Omv045F Forward Removing BamHI site in {<fliD!} cgttaagcggCatccgtgatgcc | Omv045F Forward Removing BamHI site in {<fliD!} cgttaagcggCatccgtgatgcc | ||
Omv045R Reverse Removing BamHI site in {<fliD!} ggcatcacggatGccgcttaacg | Omv045R Reverse Removing BamHI site in {<fliD!} ggcatcacggatGccgcttaacg | ||
</pre> | </pre> |
Revision as of 17:40, 10 February 2009
Madhvi Venkatesh 14:54, 2 February 2009 (EST)
Today, I got my project ball and I am making a new flagellar display system. I need to consider several possibilities and figure out what would be best to make and the source. The project description if on this page: new flagellar display system
Madhvi Venkatesh 15:40, 9 February 2009 (EST)
Possible places to find EtpA homolog (from CTEC):
APEC 01:k1 CFT073 EHEC 0157:H7 Shigella flexneri
Madhvi Venkatesh 18:21, 10 February 2009 (EST)
Construction files for C and N terminal fliD parts: Note: fliD has an internal BamHI site
Construction of {<fliD!} (N-terminal) PCR Omv044F/Omv045R on MG1655 genomic DNA (485bp, gp=A) PCR Omv045F/Omv044R on MG1655 genomic DNA (973bp, gp=B) ---- PCR Omv044F/Omv044R on A + B (1435bp, EcoRI/BamHI) Sub into pBca9495KC-Bca1144#5 (EcoRI/BamHI, 2974+910, L) Product is pBca9495KC-M10000 {<fliD!} ---- Omv044F Forward Bglbricking of {<fliD!} ccagtGAATTCatgAGATCTgcaagtatttcatcgctggg Omv044R Reverse Bglbricking of {<fliD!} gcagtGGATCCttacttggaattactgttgttttcg Omv045F Forward Removing BamHI site in {<fliD!} cgttaagcggCatccgtgatgcc Omv045R Reverse Removing BamHI site in {<fliD!} ggcatcacggatGccgcttaacg
Construction of {<fliD>} (C-terminal type 1) PCR Omv044F/Omv045R on MG1655 genomic DNA (485bp, gp=A) PCR Omv045F/Omv046R on MG1655 genomic DNA (970bp, gp=B) ---- PCR Omv044F/Omv046R on A + B (1432bp, EcoRI/BamHI) Sub into pBca9495CA-Bca1144#5 (EcoRI/BamHI, 3039+910, L) Product is pBca9495KC-M10001 {<fliD>} ---- Omv044F Forward Bglbricking of {<fliD!} ccagtGAATTCatgAGATCTgcaagtatttcatcgctggg Omv046R Reverse Bglbricking of {<fliD>} gcagtGGATCCcttggaattactgttgttttcg Omv045F Forward Removing BamHI site in {<fliD!} cgttaagcggCatccgtgatgcc Omv045R Reverse Removing BamHI site in {<fliD!} ggcatcacggatGccgcttaacg
Construction of {a~fliD>} (C-terminal type 2) PCR Omv044F/Omv045R on MG1655 genomic DNA (492bp, gp=A) PCR Omv045F/Omv046R on MG1655 genomic DNA (970bp, gp=B) ---- PCR Omv044F/Omv046R on A + B (1439bp, EcoRI/BamHI) Sub into pBca9495CA-Bca1144#5 (EcoRI/BamHI, 3039+910, L) Product is pBca9495KC-M10001 {a~fliD>} ---- Omv047F Forward Bglbricking of {a~fliD!} ccagtGAATTCatgAGATCTtcgcatggcaagtatttcatcgctggg Omv046R Reverse Bglbricking of {<fliD>} gcagtGGATCCcttggaattactgttgttttcg Omv045F Forward Removing BamHI site in {<fliD!} cgttaagcggCatccgtgatgcc Omv045R Reverse Removing BamHI site in {<fliD!} ggcatcacggatGccgcttaacg