pKLAC1

Integrative expression vector for the yeast Kluyveromyces lactis. For an alternative MCS, use pKLAC2.

Sequence Author: New England Biolabs

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HomePlasmidsYeast PlasmidspKLAC1
SnaBI (8798) BstXI (7523) SacII (7462) AanI (7195) AhdI (5918) SacII (4635) HindIII (1) PaeR7I - XhoI (257) BglII (290) AvrII (296) Acc65I (302) KpnI (306) SalI - SgrDI (308) AscI (315) NotI (324) PacI (335) StuI (340) XcmI (499) BsrGI (994) PshAI (2213) EcoO109I - PpuMI (2521) KasI (2598) NarI * (2599) SfoI (2600) BbeI (2602) Eco53kI (2926) SacI (2928) AleI (3116) BstEII (3130) AgeI (3241) TspMI - XmaI (3344) SmaI (3346) NruI (3367) Bsu36I (3388) PfoI * (3440) XbaI * (3522) AanI (3665) SbfI (3859) BmgBI (4243) BstXI (4310) PflFI - Tth111I (4353) pKLAC1 9091 bp
SnaBI  (8798)
1 site		 T A C G T A A T G C A T
BstXI  (7523)
2 sites		 C C A N N N N N N T G G G G T N N N N N N A C C
Sticky ends from different BstXI sites may not be compatible.
SacII  (7462)
2 sites		 C C G C G G G G C G C C
Efficient cleavage requires at least two copies of the SacII recognition sequence.
AanI  (7195)
2 sites		 T T A T A A A A T A T T
AhdI  (5918)
1 site		 G A C N N N N N G T C C T G N N N N N C A G
The 1-base overhangs produced by AhdI may be hard to ligate.
Sticky ends from different AhdI sites may not be compatible.
SacII  (4635)
2 sites		 C C G C G G G G C G C C
Efficient cleavage requires at least two copies of the SacII recognition sequence.
HindIII  (1)
1 site		 A A G C T T T T C G A A
PaeR7I  (257)
1 site		 C T C G A G G A G C T C
PaeR7I does not recognize the sequence CTCTCGAG.
XhoI  (257)
1 site		 C T C G A G G A G C T C
BglII  (290)
1 site		 A G A T C T T C T A G A
AvrII  (296)
1 site		 C C T A G G G G A T C C
Acc65I  (302)
1 site		 G G T A C C C C A T G G
KpnI  (306)
1 site		 G G T A C C C C A T G G
SalI  (308)
1 site		 G T C G A C C A G C T G
SgrDI  (308)
1 site		 C G T C G A C G G C A G C T G C
AscI  (315)
1 site		 G G C G C G C C C C G C G C G G
NotI  (324)
1 site		 G C G G C C G C C G C C G G C G
PacI  (335)
1 site		 T T A A T T A A A A T T A A T T
StuI  (340)
1 site		 A G G C C T T C C G G A
XcmI  (499)
1 site		 C C A N N N N N N N N N T G G G G T N N N N N N N N N A C C
The 1-base overhangs produced by XcmI may be hard to ligate.
Sticky ends from different XcmI sites may not be compatible.
BsrGI  (994)
1 site		 T G T A C A A C A T G T
BsrGI is typically used at 37°C, but is even more active at 60°C.
PshAI  (2213)
1 site		 G A C N N N N G T C C T G N N N N C A G
PshAI quickly loses activity at 37°C, but can be used at 25°C for long incubations.
EcoO109I  (2521)
1 site		 R G G N C C Y Y C C N G G R
Sticky ends from different EcoO109I sites may not be compatible.
PpuMI  (2521)
1 site		 R G G W C C Y Y C C W G G R
Sticky ends from different PpuMI sites may not be compatible.
KasI  (2598)
1 site		 G G C G C C C C G C G G
NarI  (2599)
1 site		 G G C G C C C C G C G G
* Blocked by Dcm methylation.
Efficient cleavage requires at least two copies of the NarI recognition sequence.
SfoI  (2600)
1 site		 G G C G C C C C G C G G
BbeI  (2602)
1 site		 G G C G C C C C G C G G
Cleavage may be enhanced when more than one copy of the BbeI recognition sequence is present.
Eco53kI  (2926)
1 site		 G A G C T C C T C G A G
SacI  (2928)
1 site		 G A G C T C C T C G A G
AleI  (3116)
1 site		 C A C N N N N G T G G T G N N N N C A C
BstEII  (3130)
1 site		 G G T N A C C C C A N T G G
Sticky ends from different BstEII sites may not be compatible.
BstEII is typically used at 60°C, but is 50% active at 37°C.
AgeI  (3241)
1 site		 A C C G G T T G G C C A
TspMI  (3344)
1 site		 C C C G G G G G G C C C
XmaI  (3344)
1 site		 C C C G G G G G G C C C
Cleavage may be enhanced when more than one copy of the XmaI recognition sequence is present.
SmaI  (3346)
1 site		 C C C G G G G G G C C C
SmaI can be used at 37°C for brief incubations.
NruI  (3367)
1 site		 T C G C G A A G C G C T
Bsu36I  (3388)
1 site		 C C T N A G G G G A N T C C
Sticky ends from different Bsu36I sites may not be compatible.
PfoI  (3440)
1 site		 T C C N G G A A G G N C C T
* Blocked by Dcm methylation.
Sticky ends from different PfoI sites may not be compatible.
XbaI  (3522)
1 site		 T C T A G A A G A T C T
* Blocked by Dam methylation.
AanI  (3665)
2 sites		 T T A T A A A A T A T T
SbfI  (3859)
1 site		 C C T G C A G G G G A C G T C C
BmgBI  (4243)
1 site		 C A C G T C G T G C A G
This recognition sequence is asymmetric, so ligating blunt ends generated by BmgBI will not always regenerate a BmgBI site.
BstXI  (4310)
2 sites		 C C A N N N N N N T G G G G T N N N N N N A C C
Sticky ends from different BstXI sites may not be compatible.
PflFI  (4353)
1 site		 G A C N N N G T C C T G N N N C A G
The 1-base overhangs produced by PflFI may be hard to ligate.
Sticky ends from different PflFI sites may not be compatible.
Tth111I  (4353)
1 site		 G A C N N N G T C C T G N N N C A G
The 1-base overhangs produced by Tth111I may be hard to ligate.
Sticky ends from different Tth111I sites may not be compatible.
amdS
1697 .. 3343  =  1647 bp
548 amino acids  =  60.2 kDa
Product: acetamidase from Aspergillus nidulans
confers ability to process acetamide for selection in fungal cells
amdS
1697 .. 3343  =  1647 bp
548 amino acids  =  60.2 kDa
Product: acetamidase from Aspergillus nidulans
confers ability to process acetamide for selection in fungal cells
LAC4 promoter (3')
7463 .. 8991  =  1529 bp
3' portion of a modified Kluyveromyces lactis LAC4 promoter
LAC4 promoter (3')
7463 .. 8991  =  1529 bp
3' portion of a modified Kluyveromyces lactis LAC4 promoter
AmpR
5845 .. 6705  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 2:  
   5845 .. 6636  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
5845 .. 6705  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 1:  signal sequence  
   6637 .. 6705  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
5845 .. 6705  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
ADH1 promoter
974 .. 1676  =  703 bp
promoter for alcohol dehydrogenase 1
ADH1 promoter
974 .. 1676  =  703 bp
promoter for alcohol dehydrogenase 1
LAC4 promoter (5')
3979 .. 4635  =  657 bp
5' portion of a modified Kluyveromyces lactis LAC4 promoter
LAC4 promoter (5')
3979 .. 4635  =  657 bp
5' portion of a modified Kluyveromyces lactis LAC4 promoter
LAC4 terminator
355 .. 973  =  619 bp
transcription terminator for Kluyveromyces lactis LAC4
LAC4 terminator
355 .. 973  =  619 bp
transcription terminator for Kluyveromyces lactis LAC4
ori
5086 .. 5674  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
5086 .. 5674  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
f1 ori
6837 .. 7292  =  456 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
f1 ori
6837 .. 7292  =  456 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
α-factor secretion signal
14 .. 280  =  267 bp
89 amino acids  =  9.4 kDa
Product: N-terminal secretion signal from Kluyveromyces lactis alpha-factor
Cleavage by the Kex2 protease occurs after the dibasic KR sequence. The EA dipeptides are then removed by dipeptidyl aminopeptidase A.
α-factor secretion signal
14 .. 280  =  267 bp
89 amino acids  =  9.4 kDa
Product: N-terminal secretion signal from Kluyveromyces lactis alpha-factor
Cleavage by the Kex2 protease occurs after the dibasic KR sequence. The EA dipeptides are then removed by dipeptidyl aminopeptidase A.
AmpR promoter
6706 .. 6810  =  105 bp
AmpR promoter
6706 .. 6810  =  105 bp
MCS
257 .. 343  =  87 bp
multiple cloning site
MCS
257 .. 343  =  87 bp
multiple cloning site
ORF:  2563 .. 2820  =  258 bp
ORF:  85 amino acids  =  10.2 kDa
ORF:  4435 .. 4671  =  237 bp
ORF:  78 amino acids  =  8.9 kDa
ORF:  14 .. 334  =  321 bp
ORF:  106 amino acids  =  11.1 kDa
ORF:  1697 .. 3343  =  1647 bp
ORF:  548 amino acids  =  60.2 kDa
ORF:  5975 .. 6241  =  267 bp
ORF:  88 amino acids  =  9.2 kDa
ORF:  2430 .. 2756  =  327 bp
ORF:  108 amino acids  =  11.6 kDa
ORF:  2417 .. 2806  =  390 bp
ORF:  129 amino acids  =  14.2 kDa
ORF:  2888 .. 3235  =  348 bp
ORF:  115 amino acids  =  13.5 kDa
ORF:  8939 .. 225  =  378 bp
ORF:  125 amino acids  =  14.1 kDa
ORF:  3076 .. 3474  =  399 bp
ORF:  132 amino acids  =  13.8 kDa
ORF:  5845 .. 6705  =  861 bp
ORF:  286 amino acids  =  31.6 kDa
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Download pKLAC1.dna file

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Individual Sequences & Maps

bleMX6
pFA6a-kanMX6
pPIC6(alpha) B
pYES2.1-E
hphMX6
pFA6a-kanMX6-PGAL1
pPIC6(alpha) C
pYES2 CT
kanMX
pFA6a-kanMX6-PGAL1-3HA
pPIC9
pYES2 NT A
kanMX2
pFA6a-kanMX6-PGAL1-GFP
pPIC9K
pYES2 NT B
kanMX4
pFA6a-kanMX6-PGAL1-GST
pPICZ A
pYES2 NT C
kanMX6
pFA6a-natMX6
pPICZ B
pYES3 CT
natMX6
pFA6a-TRP1
pPICZ C
pYES6 CT
pAbAi
pFA6a-TRP1-PGAL1
pPICZ(alpha) A
pYM25
pAD-GAL4
pFA6a-TRP1-PGAL1-3HA
pPICZ(alpha) B
pYM26
pAG25
pFA6a-TRP1-PGAL1-GFP
pPICZ(alpha) C
pYM44
pAG29
pFA6a-TRP1-PGAL1-GST
pPink-HC
YCp50
pAG32
pFLD
pPink-LC
YCp50-poly
pAO815
pFLD(alpha)
pPINK(alpha)-HC
YCplac111
patMX4
pGAD-C1
pRS303
YCplac22
pAUR101
pGAD-C2
pRS304
YCplac33
pAUR112
pGAD-C3
pRS305
YEp13
pAUR123
pGADT7 AD
pRS306
YEp24
pAUR224
pGADT7-Rec
pRS313
YEp351
pBridge
pGADT7-Rec (linearized)
pRS314
YEp352
pDH25
pGAPZ A
pRS315
YEp353
pEG202
pGAPZ B
pRS316
YEp354
pESC-HIS
pGAPZ C
pRS403
YEp355
pESC-LEU
pGAPZ(alpha) A
pRS404
YEp356
pESC-TRP
pGAPZ(alpha) B
pRS405
YEp356R
pESC-URA
pGAPZ(alpha) C
pRS406
YEp357
pFA6-kanMX4
pGBD-C1
pRS413
YEp357R
pFA6a
pGBD-C2
pRS414
YEp358
pFA6a-12Pk-kanMX6
pGBD-C3
pRS415
YEp358R
pFA6a-13Myc-His3MX6
pGBDU-C1
pRS416
YEp363
pFA6a-13Myc-kanMX6
pGBDU-C2
pRS423
YEp364
pFA6a-13Myc-TRP1
pGBDU-C3
pRS424
YEp365
pFA6a-3HA-His3MX6
pGBKT7
pRS425
YEp366
pFA6a-3HA-kanMX6
pGilda
pRS426
YEp366R
pFA6a-3HA-TRP1
pHIL-D2
pSpot5
YEp367
pFA6a-5FLAG-kanMX6
pHIL-S1
pSpot6
YEp367R
pFA6a-bleMX6
pJG4-5
pSpot7
YEp368
pFA6a-GFP(S65T)-His3MX6
pKLAC1
pSpot8
YEp368R
pFA6a-GFP(S65T)-kanMX6
pKLAC2
pTEF1 Zeo
YEplac112
pFA6a-GFP(S65T)-TRP1
pOAD
pUCATPH
YEplac181
pFA6a-GST-His3MX6
pOBD2
pY-secSUMOstar
YEplac195
pFA6a-GST-kanMX6
pP-secSUMO3
pY-SUMOstar
YIp351
pFA6a-GST-TRP1
pPIC3
pYC2-E
YIp352
pFA6a-His3MX6
pPIC3.5
pYC2 CT
YIp5
pFA6a-His3MX6-PGAL1
pPIC3.5K
pYC2 NT A
YIplac128
pFA6a-His3MX6-PGAL1-3HA
pPIC6 A
pYC2 NT B
YIplac204
pFA6a-His3MX6-PGAL1-GFP
pPIC6 B
pYC2 NT C
YIplac211
pFA6a-His3MX6-PGAL1-GST
pPIC6 C
pYC6 CT
YRp7
pFA6a-hphMX6
pPIC6(alpha) A
pYES2
alpha-factor secretion signal

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