pHELLSGATE 12

High-throughput binary Gateway® destination vector with two introns, for silencing genes in plants using intron-containing hairpin RNA.
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AgeI (17,247) XbaI (16,903) XbaI (15,484) BspDI - ClaI (15,262) PacI (15,151) PDK intron KpnI (14,465) Acc65I (14,461) EcoRI (14,455) XhoI (14,449) XhoI (13,020) ScaI (12,386) StuI (12,234) SacI (11,646) Eco53kI (11,644) PmeI (11,220) BsiWI (11,055) FseI (10,595) Bsu36I (330) RsrII (1119) AflII (1923) MauBI (2118) LB T-DNA repeat BspEI (3460) AleI (5445) SexAI * (5533) FspAI (6101) NruI (6598) pHELLSGATE 12 17,681 bp
AgeI  (17,247)
1 site
A C C G G T T G G C C A
XbaI  (16,903)
2 sites
T C T A G A A G A T C T
XbaI  (15,484)
2 sites
T C T A G A A G A T C T
BspDI  (15,262)
1 site
A T C G A T T A G C T A
ClaI  (15,262)
1 site
A T C G A T T A G C T A
PacI  (15,151)
1 site
T T A A T T A A A A T T A A T T
KpnI  (14,465)
1 site
G G T A C C C C A T G G
Acc65I  (14,461)
1 site
G G T A C C C C A T G G
EcoRI  (14,455)
1 site
G A A T T C C T T A A G
XhoI  (14,449)
2 sites
C T C G A G G A G C T C
XhoI  (13,020)
2 sites
C T C G A G G A G C T C
ScaI  (12,386)
1 site
A G T A C T T C A T G A
StuI  (12,234)
1 site
A G G C C T T C C G G A
SacI  (11,646)
1 site
G A G C T C C T C G A G
Eco53kI  (11,644)
1 site
G A G C T C C T C G A G
PmeI  (11,220)
1 site
G T T T A A A C C A A A T T T G
BsiWI  (11,055)
1 site
C G T A C G G C A T G C

BsiWI is typically used at 55°C, but is 50% active at 37°C.
FseI  (10,595)
1 site
G G C C G G C C C C G G C C G G

FseI gradually loses activity when stored at -20°C.
Bsu36I  (330)
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.
RsrII  (1119)
1 site
C G G W C C G G C C W G G C

Efficient cleavage requires at least two copies of the RsrII recognition sequence.
Sticky ends from different RsrII sites may not be compatible.
For full activity, add fresh DTT.
AflII  (1923)
1 site
C T T A A G G A A T T C
MauBI  (2118)
1 site
C G C G C G C G G C G C G C G C
BspEI  (3460)
1 site
T C C G G A A G G C