Addgene - pKJ1267 (aka pAC-alphaGal4) Plasmid Data

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Keith Joung Lab Plasmids

This is commonly
requested with

pKJ1712

Strain KJ1C

Strain CSH100

pMLM36 (aka pBR-UV5-GP- FD2)

pAC-Kan-alphaGal4

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Plasmid 21869: pKJ1267 (aka pAC-alphaGal4)

Gene/insert name:		None
Insert size (bp):		Unknown
Vector backbone:		pKJ1267 (Search Vector Database)
Type of vector:		Bacterial expression
Backbone size (bp):		3428
5' Sequencing primer:		lac promoter (List of Sequencing Primers)
Bacteria resistance:		Chloramphenicol
High or low copy:		Low Copy
Grow in standard E. coli @ 37C:		No
Please specify bacterial strain for growth and growth condition:		Grow in LB supplemented with 30 µg/ml chloramphenicol. Due to toxicity associated with high expression from the dual lpp/lacUV5 promoter, this plasmid must be propagated in a lacIq strain (e.g.--XL-1 Blue).
Sequence:		View sequence
Plasmid Provided In:		XL1 Blue
Principal Investigator:		Keith Joung
Terms and Licenses:		MTA

Comments: Bacterial expression vector for the alpha-Gal4 hybrid protein (required for the B2H system). The tandem lpp/lacUV5 promoter which drives expression of the Gal11P fusion protein contains a lac operator and therefore these dual promoters can be repressed by lac repressor.

This plasmid is a very low copy number plasmid and therefore we recommend that maxipreps be performed from a 200 ml culture. A miniprep will not be sufficient.

Annotations for pKJ1267:

2988- 219: Chloramphenicol-resistance gene (complementary strand)/ 581-1493: p15A origin of replication/ 1552-1557: lpp promoter -35 box / 1576-1581: lpp promoter -10 box / 1624-1629 : lacUV5 promoter -35 box / 1648-1653 : lacUV5 promoter -10 box / 1748-2623 : alpha-Gal4 hybrid protein gene (E. coli RNA polymerase alpha subunit aa 1-248 fused by an Ala-Ala-Ala linker to aa 58-97 of the yeast Gal4 protein)

Addgene has sequenced a portion of this plasmid for verification. Click here for the sequencing result.

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Selected features

ORF frame 3

219 - 2988

CAT/CamR

219 - 2988

CAT/CamR(660-444)

219 - 4

p15A_origin

581 - 1493

lac_promoter

1624 - 1647

M13_pUC_rev_primer

1667 - 1689

M13_reverse_primer

1688 - 1706

ORF frame 2

1748 - 2623

Unique restriction sites

NheI		583
SacII		836
AseI		1407
PstI		2083
PmlI		2131
SalI		2264
NotI		2493
XhoI		2547
HpaI		2607
BglII		2624
BclI		2725
NcoI		3128

Please acknowledge the principal investigator if you use this plasmid in a publication.

Also, please include the text "Addgene plasmid 21869" in your Materials and Methods section. This information allows Addgene to create a link from the plasmid page to your publication.