pRS416

Yeast centromere vector with a URA3 marker and an MCS derived from pBLUESCRIPT II.
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HomePlasmidsYeast PlasmidspRS416
SwaI (4354) PmlI (4330) BsaHI (3934) NmeAIII (3545) AhdI (3397) PfoI (46) NdeI (330) BsgI (363) SbfI (399) BfuAI - BspMI (402) XcmI (545) NcoI (622) BstBI (682) BsmI (779) StuI (853) Bpu10I (1147) NsiI (1244) BtgZI (1576) NgoMIV (1676) NaeI (1678) Acc65I (2004) KpnI (2008) AbsI - PaeR7I - PspXI - XhoI (2019) SalI (2025) BspDI - ClaI (2035) HindIII (2040) EcoRI (2052) TspMI - XmaI (2064) SmaI (2066) BamHI (2070) SpeI (2076) XbaI (2082) EagI - NotI (2089) AleI (2100) SacII (2101) BstXI (2102) Eco53kI (2108) SacI (2110) lac operator pRS416 4898 bp
SwaI  (4354)
1 site		 A T T T A A A T T A A A T T T A
SwaI is typically used at 25°C, but is 50% active at 37°C.
PmlI  (4330)
1 site		 C A C G T G G T G C A C
BsaHI  (3934)
1 site		 G R C G Y C C Y G C R G
BsaHI is typically used at 37°C, but is even more active at 60°C.
NmeAIII  (3545)
1 site		 G C C G A G ( N ) 18-19 N N C G G C T C ( N ) 18-19
Efficient cleavage requires at least two copies of the NmeAIII recognition sequence.
Sticky ends from different NmeAIII sites may not be compatible.
For full activity, add fresh S-adenosylmethionine (SAM).
AhdI  (3397)
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.
PfoI  (46)
1 site		 T C C N G G A A G G N C C T
Sticky ends from different PfoI sites may not be compatible.
NdeI  (330)
1 site		 C A T A T G G T A T A C
Prolonged incubation with NdeI may lead to removal of additional nucleotides.
BsgI  (363)
1 site		 G T G C A G ( N ) 14 N N C A C G T C ( N ) 14
Efficient cleavage requires at least two copies of the BsgI recognition sequence.
Sticky ends from different BsgI sites may not be compatible.
For full activity, add fresh S-adenosylmethionine (SAM).
SbfI  (399)
1 site		 C C T G C A G G G G A C G T C C
BfuAI  (402)
1 site		 A C C T G C ( N ) 4 T G G A C G ( N ) 4 ( N ) 4
Efficient cleavage requires at least two copies of the BfuAI recognition sequence.
Sticky ends from different BfuAI sites may not be compatible.
BfuAI is typically used at 50°C, but is 50% active at 37°C.
BspMI  (402)
1 site		 A C C T G C ( N ) 4 T G G A C G ( N ) 4 ( N ) 4
Efficient cleavage requires at least two copies of the BspMI recognition sequence.
Sticky ends from different BspMI sites may not be compatible.
XcmI  (545)
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.
NcoI  (622)
1 site		 C C A T G G G G T A C C
BstBI  (682)
1 site		 T T C G A A A A G C T T
BsmI  (779)
1 site		 G A A T G C N C T T A C G N
Sticky ends from different BsmI sites may not be compatible.
StuI  (853)
1 site		 A G G C C T T C C G G A
Bpu10I  (1147)
1 site		 C C T N A G C G G A N T C G
Cleavage may be enhanced when more than one copy of the Bpu10I recognition sequence is present.
This recognition sequence is asymmetric, so ligating sticky ends generated by Bpu10I will not always regenerate a Bpu10I site.
Sticky ends from different Bpu10I sites may not be compatible.
NsiI  (1244)
1 site		 A T G C A T T A C G T A
BtgZI  (1576)
1 site		 G C G A T G ( N ) 10 C G C T A C ( N ) 10 ( N ) 4
Sticky ends from different BtgZI sites may not be compatible.
After cleavage, BtgZI can remain bound to DNA and alter its electrophoretic mobility.
BtgZI is typically used at 60°C, but is 75% active at 37°C.
NgoMIV  (1676)
1 site		 G C C G G C C G G C C G
Efficient cleavage requires at least two copies of the NgoMIV recognition sequence.
NaeI  (1678)
1 site		 G C C G G C C G G C C G
Efficient cleavage requires at least two copies of the NaeI recognition sequence.
Acc65I  (2004)
1 site		 G G T A C C C C A T G G
KpnI  (2008)
1 site		 G G T A C C C C A T G G
AbsI  (2019)
1 site		 C C T C G A G G G G A G C T C C
PaeR7I  (2019)
1 site		 C T C G A G G A G C T C
PaeR7I does not recognize the sequence CTCTCGAG.
PspXI  (2019)
1 site		 V C T C G A G B B G A G C T C V
XhoI  (2019)
1 site		 C T C G A G G A G C T C
SalI  (2025)
1 site		 G T C G A C C A G C T G
BspDI  (2035)
1 site		 A T C G A T T A G C T A
ClaI  (2035)
1 site		 A T C G A T T A G C T A
HindIII  (2040)
1 site		 A A G C T T T T C G A A
EcoRI  (2052)
1 site		 G A A T T C C T T A A G
TspMI  (2064)
1 site		 C C C G G G G G G C C C
XmaI  (2064)
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  (2066)
1 site		 C C C G G G G G G C C C
SmaI can be used at 37°C for brief incubations.
BamHI  (2070)
1 site		 G G A T C C C C T A G G
After cleavage, BamHI-HF® (but not the original BamHI) can remain bound to DNA and alter its electrophoretic mobility.
SpeI  (2076)
1 site		 A C T A G T T G A T C A
XbaI  (2082)
1 site		 T C T A G A A G A T C T
EagI  (2089)
1 site		 C G G C C G G C C G G C
NotI  (2089)
1 site		 G C G G C C G C C G C C G G C G
AleI  (2100)
1 site		 C A C N N N N G T G G T G N N N N C A C
SacII  (2101)
1 site		 C C G C G G G G C G C C
Efficient cleavage requires at least two copies of the SacII recognition sequence.
BstXI  (2102)
1 site		 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.
Eco53kI  (2108)
1 site		 G A G C T C C T C G A G
SacI  (2110)
1 site		 G A G C T C C T C G A G
AmpR
3324 .. 4184  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 2:  
   3324 .. 4115  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
3324 .. 4184  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 1:  signal sequence  
   4116 .. 4184  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
3324 .. 4184  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
URA3
417 .. 1220  =  804 bp
267 amino acids  =  29.3 kDa
Product: orotidine-5'-phosphate decarboxylase, required for uracil biosynthesis
yeast auxotrophic marker, counterselectable with 5-fluoroorotic acid (5-FOA)
URA3
417 .. 1220  =  804 bp
267 amino acids  =  29.3 kDa
Product: orotidine-5'-phosphate decarboxylase, required for uracil biosynthesis
yeast auxotrophic marker, counterselectable with 5-fluoroorotic acid (5-FOA)
ori
2565 .. 3153  =  589 bp
high-copy-number colE1/pMB1/pBR322/pUC origin of replication
ori
2565 .. 3153  =  589 bp
high-copy-number colE1/pMB1/pBR322/pUC origin of replication
lacZα
1589 .. 2167  =  579 bp
192 amino acids  =  20.6 kDa
Product: LacZα fragment of β-galactosidase
lacZα
1589 .. 2167  =  579 bp
192 amino acids  =  20.6 kDa
Product: LacZα fragment of β-galactosidase
CEN/ARS
4326 .. 4829  =  504 bp
S. cerevisiae CEN6 centromere fused to an autonomously replicating sequence
CEN/ARS
4326 .. 4829  =  504 bp
S. cerevisiae CEN6 centromere fused to an autonomously replicating sequence
URA3 promoter
196 .. 416  =  221 bp
URA3 promoter
196 .. 416  =  221 bp
AmpR promoter
4185 .. 4289  =  105 bp
AmpR promoter
4185 .. 4289  =  105 bp
lac promoter
2211 .. 2241  =  31 bp
3 segments
   Segment 3:  -10  
   2211 .. 2217  =  7 bp
promoter for the E. coli lac operon
lac promoter
2211 .. 2241  =  31 bp
3 segments
   Segment 2:  
   2218 .. 2235  =  18 bp
promoter for the E. coli lac operon
lac promoter
2211 .. 2241  =  31 bp
3 segments
   Segment 1:  -35  
   2236 .. 2241  =  6 bp
promoter for the E. coli lac operon
lac promoter
2211 .. 2241  =  31 bp
3 segments
promoter for the E. coli lac operon
lac operator
2187 .. 2203  =  17 bp
The lac repressor binds to the lac operator to inhibit transcription in E. coli. This inhibition can be relieved by adding lactose or isopropyl-β-D-thiogalactopyranoside (IPTG).
lac operator
2187 .. 2203  =  17 bp
The lac repressor binds to the lac operator to inhibit transcription in E. coli. This inhibition can be relieved by adding lactose or isopropyl-β-D-thiogalactopyranoside (IPTG).
f1 ori
1351 .. 1806  =  456 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
f1 ori
1351 .. 1806  =  456 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
MCS
2004 .. 2111  =  108 bp
pBluescript multiple cloning site
MCS
2004 .. 2111  =  108 bp
pBluescript multiple cloning site
T7 promoter
1977 .. 1995  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T7 promoter
1977 .. 1995  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T3 promoter
2124 .. 2142  =  19 bp
promoter for bacteriophage T3 RNA polymerase
T3 promoter
2124 .. 2142  =  19 bp
promoter for bacteriophage T3 RNA polymerase
M13 fwd
1951 .. 1967  =  17 bp
common sequencing primer, one of multiple similar variants
M13 fwd
1951 .. 1967  =  17 bp
common sequencing primer, one of multiple similar variants
M13 rev
2163 .. 2179  =  17 bp
common sequencing primer, one of multiple similar variants
M13 rev
2163 .. 2179  =  17 bp
common sequencing primer, one of multiple similar variants
ORF:  3454 .. 3720  =  267 bp
ORF:  88 amino acids  =  9.2 kDa
ORF:  417 .. 1220  =  804 bp
ORF:  267 amino acids  =  29.3 kDa
ORF:  3324 .. 4184  =  861 bp
ORF:  286 amino acids  =  31.6 kDa
ORF:  452 .. 928  =  477 bp
ORF:  158 amino acids  =  16.8 kDa
ORF:  1589 .. 2167  =  579 bp
ORF:  192 amino acids  =  20.6 kDa
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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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