Team:Paris/Freezer

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Revision as of 16:26, 4 August 2009

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Freezer

Color code for sticker
- Vector
- Insert
- PCR product
- Plasmid
- Kanamycin
- Ampicilin
- Chloranphenicol

Primers

[O1] Rev P1
Description	tolR
Sequence (5' > 3')	TCTGCTGCAGCGGCCGCTACTAGTATTAGTAAGGCACATCTTTTGCGCCACCG
Designer	D. Bikard
Worked

[O2] Fwd P2
Description	tolR (DO NOT USE ERROR)
Sequence (5' > 3')	TCTGGAATTCGCGGCCGCTTCTAGATGAAATCAACATTGTACCGTTGCTGGACGTACTG
Designer	D. Bikard
Worked

[O3] Rev P3
Description	tolA
Sequence (5' > 3')	TCTGCTGCAGCGGCCGCTACTAGTATTATTACGGTTTGAAGTCCAATGGCG
Designer	D. Bikard
Worked

[O4] Rev P4
Description	tolA (ERROR: Not designed for N-term fusion)
Sequence (5' > 3')	TCTGGAATTCGCGGCCGCTTCTAGAATGGGTAATACTAAAAACAATGGCGC
Designer	D. Bikard
Worked

[O5] Rev P5
Description	tatABC
Sequence (5' > 3')	TCTGCTGCAGCGGCCGCTACTAGTATTATTATTCTTCAGTTTTTTCGCTTTCTGC
Designer	D. Bikard
Worked

[06] Fwd P6
Description	tatABC (without RBS)
Sequence (5' > 3')	TCTGGAATTCGCGGCCGCTTCTAGAATGGGTGGTATCAGTATTTGG
Designer	D. Bikard
Worked

[O7] Rev P7
Description	clyA fusion
Sequence (5' > 3')	TTTCTGCAGCGGCCGCTACTAGTGACTTCAGGTACCTCAAAGAGTGTC
Designer	D. Bikard
Worked

[O8] Fwd P8
Description	clyA fusion (ERROR: No poly-Gly linker)
Sequence (5' > 3')	TTTGAATTCGCGGCCGCTTCTAGAATGACTGAAATCGTTGCAGATAAAA
Designer	D. Bikard
Worked

[O9] clyA_Rw
Description	clyA
Sequence (5' > 3')	GTTTCTTCCTGCAGCGGCCGCTACTAGTATTATTAGACTTCAGGTACCTCAAAGAG
Designer	G. Cambray
Not Tested

[O10] clyA_fusion_polyG_Rw
Description	clyA fusion (with 5-Gly linker)
Sequence (5' > 3')	GTTTCTTCCTGCAGCGGCCGCTACTAGTGCCGCCGCCGCCGCCGACTTCAGGTACCTCAAAGAG
Designer	G. Cambray
Not Tested

[O11] ompA_signal_Fw
Description	ompA signal for export to periplasm - from expression vector RBS as in pIN-III-ompA-2 vector - keeps vector original RBS
Sequence (5' > 3')	GTTTCTTCGAATTCGCGGCCGCTTCTAGAGAACGAGGGCAAAAAATGAAAAAG
Designer	G. Cambray
Not Tested

[O12] ompA_signal_fusion_Rw
Description	ompA signal for export to periplasm - from expression vector pIN-III-ompA-2 - original RBS from the vector
Sequence (5' > 3')	GTTTCTTCCTGCAGCGGCCGCTACTAGTGGCCTGCGCTACGGTAGCGAAAC
Designer	G. Cambray
Not Tested

[O13] tolR_II_Fusion_Fw
Description	For fusion of tolR domain II (standard 23)
Sequence (5' > 3')	GTTTCTTCGAATTCGCGGCCGCTTCTAGAGAGGTCGATCTGCCAGACGCTAC
Designer	G. Cambray
Not Tested

[O14] tatABC_RBS_Fw
Description	Amplification from tatA - contains RBS
Sequence (5' > 3')	TTGAATTCGCGGCCGCTTCTAGAAGGAGGTATATAATGATGGGTGGTATCAGTATTTGG
Designer	G. Cambray
Not Tested

[O15] TE3_fusion_Fw
Description	Fusion of N-term domain of colicin E3 (standard 23)
Sequence (5' > 3')	GTTTCTTCGAATTCGCGGCCGCTTCTAGAAGCGGTGGCGATGGACGCGGCCATAAC
Designer	G. Cambray
Not Tested

[O16] TE3_Rw
Description	Amplification of N-term domain of colicin E3 (standard 23)
Sequence (5' > 3')	GTTTCTTCCTGCAGCGGCCGCTACTAGTATTATTAGCGTTCATAATTTCGCTCAG
Designer	G. Cambray
Not Tested

[O17] Tg3p_Rw
Description	Amplification of N-term domain of M13's g3p
Sequence (5' > 3')	GTTTCTTCCTGCAGCGGCCGCTACTAGTATTATTAACACTGAGTTTCGTCACCAGTAC
Designer	G. Cambray
Not Tested

[O18] Tg3p_fusion_Fw
Description	Fusion of N-term domain of M13's g3p (standard 23)
Sequence (5' > 3')	GTTTCTTCGAATTCGCGGCCGCTTCTAGAAAAAAATTATTATTCGCAATTCCTTTAG
Designer	G. Cambray
Not Tested

[O19] g3p_F
Description	g3p
Sequence (5' > 3')	GTTTCTTCGAATTCGCGGCCGCTTCTAGAGCTCCGCTGAAACTGTTGAAAGTTG
Designer	G. Cambray
Not Tested

[O20] g3p_R
Description	g3p
Sequence (5' > 3')	GTTTCTTCCTGCAGCGGCCGCTACTAGTATTATTAAGACTCCTTATTACGCAG
Designer	G. Cambray
Not Tested

[O21] Ompa_linker_F
Description	For standardisation of OmpA linker (BBaK103006)
Sequence (5' > 3')	TTGAATTCGCGGCCGCTTCTAGAAGGAGGTATATAATGAAAGCTACTAAACTGGTACTGG
Designer	G. Cambray
Not Tested

[O22] Ompa_linker_R
Description	For standardisation of OmpA linker (BBaK103006)
Sequence (5' > 3')	GTTTCTTCCTGCAGCGGCCGCTACTAGTAGAGCTCCCTCCTCCAGAACC
Designer	G. Cambray
Not Tested

[O23] OmpAsignal-TE3_seq_Fw
Description	???
Sequence (5' > 3')	CACAAATAGCGAAAGATGAC
Designer	G. Cambray
Not Tested

[O24] OmpAsignal-TE3_seq_Rw
Description	???
Sequence (5' > 3')	ATCATCTGCGGGTAATGATG
Designer	G. Cambray
Not Tested

[O25] VAMP2_Seq_Fw
Description	???
Sequence (5' > 3')	ACTCACTATAGGGAGACCACA
Designer	G. Cambray
Not Tested

[O26] VAMP2_Seq_Rv
Description	???
Sequence (5' > 3')	GGGCTTTGTTAGCAGCCGGAT
Designer	G. Cambray
Not Tested

[O27] ClyA_seq_Fw
Description	???
Sequence (5' > 3')	GCAGCTATTTCCAGTCACAG
Designer	G. Cambray
Not Tested

[O28] ClyA_seq_Rw
Description	???
Sequence (5' > 3')	CGCCCGCAGCAATAGAATAG
Designer	G. Cambray
Not Tested

[O29] PFecA_Fw
Description	???
Sequence (5' > 3')	GTTTCTTCGAATTCGCGGCCGCTTCTAGAGTACGCGGTACTGGATAAACATTTC
Designer	G. Cambray
Not Tested

[O30] PFecA_Rv
Description	???
Sequence (5' > 3')	GTTTCTTCCTGCAGCGGCCGCTACTAGTATTTTTGTTGTGTTCAGCTATGAGT
Designer	G. Cambray
Not Tested

PCR products

PCR products
Number	Name	Oligo F	Oligo R	Tm	Matrix	Length	Comments	Quantity	[]ng/µL	Date
Number	g3p_std	[O19] g3p_F	[O20] g3p_R	Tm	Matrix	Length	Comments	?	? ng/µL	?
Number	OmpA_L_std	[O21] Ompa_linker_F	[O22] Ompa_linker_R	Tm	Matrix	Length	Comments	?	? ng/µL	?

http://parts.mit.edu/r/parts/images/partbypart/icon_terminator.png http://parts.mit.edu/images/partbypart/icon_regulatory.png http://parts.mit.edu/r/parts/images/partbypart/icon_rbs.png http://parts.mit.edu/r/parts/images/partbypart/icon_reporter.png http://parts.mit.edu/registry/images/7/7b/Part_icon_cds.png http://parts.mit.edu/registry/images/9/9f/Part_icon_dna.png http://parts.mit.edu/registry/images/2/2e/Part_icon_generator.png http://parts.mit.edu/registry/images/a/ab/Part_icon_plasmid.png

Plasmids: MiniPrep Products

Plasmids: MiniPrep Products
Number	Name	Description	Products in use	Date
P1	pSB2K3	General plasmid for cloning step after PCR	Products in use	08/03
P2	Bba-J61002	Plasmid backbone, it allows us to change promoter RFP as promoter	Products in use	08/03
P3	Bba-J32015	PelB leader sequence	Products in use	08/03
P4	Bba-B0014	Double terminator bidirectionnal (B0012-B0011)	Products in use	08/03
P5	Bba-I14032	pLac	Products in use	08/03
P6	BBa_K103006	OmpA linker (464bp)	Products in use	08/03
P7	Bba-R0040	TetR repressible promoter	Products in use	08/03
P8	Bba-I0500	pBad/araC	Products in use	08/03
P9	ΔN-Syn1A	???	Products in use	08/03
P10	His6-SNAP25	???	Products in use	08/03
P11	VAMP2-HiB6	???	Products in use	08/03
P12	g3p	g3p in pHEN2 plasmid	Products in use	08/04

Digestion Products

Red = Vector

Yellow = Insert

Digestion Products
Number	Product Name	Matrix Name	Enz. 1	Enz. 2	Size	Description	Product Left	[]ng/µL	Date
D1	P3	BBa-J32015	XbaI	PstI	Size	???	???	???	08/04
D2	P4	BBa-B0014	XbaI	PstI	Size	???	???	???	08/04
D3	P5	BBa-I14032	XbaI	PstI	Size	???	???	???	08/04
D4	P7	Bba-R0040	XbaI	PstI	Size	???	???	???	08/04
D5	P8	BBa-10500	XbaI	PstI	Size	???	???	???	08/04
D6	???	???	XbaI	PstI	Size	???	???	???	08/04
D7	???	???	XbaI	PstI	Size	???	???	???	08/04

Unexpected digestion results

Digestion Products
Number	Matrix Name	Enz. 1	Enz. 2	Expected Size	Description	Date

Strains (Glycerol stock)

Strains glycerol stock
Number	R	Name	Description	Date
S1		~~DH5α~~	~~DH5α E. Coli strain for chimically competent~~	07/20
S2	K	K12 MG 1655 ΔrecA::Kan	Standard E.Coli strain, Kan resistance	07/20
S3	K	W3110 Δnlp1	From Daniel Rodriguez, Kan resistance	07/23
TOP10		TOP10	Competent bacteria	07/27
S4		TB28	No kanamycin resistance	07/29
S5	K	MG4	Strain needs to be grown on media with [NaCl]>=0.5%, preferably 1%NaCl	07/29
S6		DH5α	DH5α E. Coli strain for chimically competent (Pasteur)	07/30
S7	A	DH5α(g3p)	DH5α E. Coli strain with g3p in plasmid, Amp resistance	07/30
S8	K	DH5α(pSB2K3)	DH5α E. Coli strain with pSB2K3 plasmid, Kan resistance	08/03
S9	A	DH5α(BBa_J61002)	DH5α E. Coli strain with BBa_J61002 in plasmid, Amp resistance	08/03
S10	K	DH5α(BBa_J32015)	DH5α E. Coli strain with BBa_J32015 in plasmid, Kan resistance	08/03
S11	K	DH5α(BBa_B0014)	DH5α E. Coli strain with BBa_B0014 in plasmid, Kan resistance	08/03
S12	K	DH5α(BBa_I14032)	DH5α E. Coli strain with BBa_I14032 in plasmid, Kan resistance	08/03
S13	K	DH5α(BBa_K103006)	DH5α E. Coli strain with BBa_K103006 in plasmid, Kan resistance	08/03
S14	A	DH5α(BBa_R0040)	DH5α E. Coli strain with BBa_R0040 in plasmid, Amp resistance	08/03
S15	K	DH5α(BBa_10500)	DH5α E. Coli strain with BBa_10500 in plasmid, Kan resistance	08/03
S16	K	DH5α(ΔN-Syn1A)	DH5α E. Coli strain with ΔN-Syn1A in plasmid, Kan resistance	08/03
S17	A	DH5α(His6-SNAP25)	DH5α E. Coli strain with His6-SNAP25 in plasmid, Amp resistance	08/03
S18	A	DH5α(Vamp2-His6)	DH5α E. Coli strain with Vamp2-His6 in plasmid, Amp resistance	08/03
S19	A	DH5α(puc19)	DH5α E. Coli strain with puc19 plasmid, Amp resistance	07/30

@@ Line 540: / Line 540: @@
 |- style="background: #ccccff; text-align: center;"
 |width=5%| Number
-|width=10%| OmpA_L_std
+|width=10%| g3p_std
-|width=10%| Oligo F
+|width=10%| [O19] g3p_F
-|width=10%| Oligo R
+|width=10%| [O20] g3p_R
 |width=5%| Tm
 |width=10%| Matrix
 |width=10%| Length
 |width=25%| Comments
-|width=5%| Quantity
+|width=5%| ?
-|width=5%| []ng/µL
+|width=5%| ? ng/µL
-|width=5%| Date
+|width=5%| ?
 |- style="background: #ccccff; text-align: center;"
 |width=5%| Number
-|width=10%| g3p_std
+|width=10%| OmpA_L_std
-|width=10%| Oligo F
+|width=10%| [O21] Ompa_linker_F
-|width=10%| Oligo R
+|width=10%| [O22] Ompa_linker_R
 |width=5%| Tm
 |width=10%| Matrix
 |width=10%| Length
 |width=25%| Comments
-|width=5%| Quantity
+|width=5%| ?
-|width=5%| []ng/µL
+|width=5%| ? ng/µL
-|width=5%| Date
+|width=5%| ?
 |}
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