pCFD1-dU6-1gRNA

Drosophila expression vector in which a guide RNA (gRNA) is expressed from the U6-1 promoter.

Sequence Author: Bullock Lab / Addgene #49408

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BglII (6619) EcoRI (1) BbsI (799) BbsI (821) polIII terminator XbaI (1132) pCFD1-dU6:1gRNA 6624 bp
BglII  (6619)
1 site
A G A T C T T C T A G A
EcoRI  (1)
1 site
G A A T T C C T T A A G
BbsI  (799)
2 sites
G A A G A C N N C T T C T G N N ( N ) 4

Sticky ends from different BbsI sites may not be compatible.
BbsI gradually loses activity when stored at -20°C.
BbsI  (821)
2 sites
G A A G A C N N C T T C T G N N ( N ) 4

Sticky ends from different BbsI sites may not be compatible.
BbsI gradually loses activity when stored at -20°C.
XbaI  (1132)
1 site
T C T A G A A G A T C T
vermilion
4229 .. 5722  =  1494 bp
379 amino acids  =  44.4 kDa
6 segments  =  1140 bp
Product: tryptophan oxygenase
selectable marker, required to synthesize the brown eye pigment in Drosophila
vermilion
4229 .. 5722  =  1494 bp
379 amino acids  =  44.4 kDa
6 segments  =  1140 bp
Product: tryptophan oxygenase
selectable marker, required to synthesize the brown eye pigment in Drosophila
vermilion
4229 .. 5722  =  1494 bp
379 amino acids  =  44.4 kDa
6 segments  =  1140 bp
Product: tryptophan oxygenase
selectable marker, required to synthesize the brown eye pigment in Drosophila
vermilion
4229 .. 5722  =  1494 bp
379 amino acids  =  44.4 kDa
6 segments  =  1140 bp
Product: tryptophan oxygenase
selectable marker, required to synthesize the brown eye pigment in Drosophila
vermilion
4229 .. 5722  =  1494 bp
379 amino acids  =  44.4 kDa
6 segments  =  1140 bp
Product: tryptophan oxygenase
selectable marker, required to synthesize the brown eye pigment in Drosophila
vermilion
4229 .. 5722  =  1494 bp
379 amino acids  =  44.4 kDa
6 segments  =  1140 bp
   Segment 6:  
   4229 .. 4346  =  118 bp
   38 amino acids  =  4.5 kDa
Product: tryptophan oxygenase
selectable marker, required to synthesize the brown eye pigment in Drosophila
vermilion
4229 .. 5722  =  1494 bp
379 amino acids  =  44.4 kDa
6 segments  =  1140 bp
   Segment 5:  
   4423 .. 4516  =  94 bp
   32 amino acids  =  3.5 kDa
Product: tryptophan oxygenase
selectable marker, required to synthesize the brown eye pigment in Drosophila
vermilion
4229 .. 5722  =  1494 bp
379 amino acids  =  44.4 kDa
6 segments  =  1140 bp
   Segment 4:  
   4608 .. 5214  =  607 bp
   202 amino acids  =  24.0 kDa
Product: tryptophan oxygenase
selectable marker, required to synthesize the brown eye pigment in Drosophila
vermilion
4229 .. 5722  =  1494 bp
379 amino acids  =  44.4 kDa
6 segments  =  1140 bp
   Segment 3:  
   5269 .. 5402  =  134 bp
   45 amino acids  =  5.5 kDa
Product: tryptophan oxygenase
selectable marker, required to synthesize the brown eye pigment in Drosophila
vermilion
4229 .. 5722  =  1494 bp
379 amino acids  =  44.4 kDa
6 segments  =  1140 bp
   Segment 2:  
   5463 .. 5623  =  161 bp
   53 amino acids  =  6.1 kDa
Product: tryptophan oxygenase
selectable marker, required to synthesize the brown eye pigment in Drosophila
vermilion
4229 .. 5722  =  1494 bp
379 amino acids  =  44.4 kDa
6 segments  =  1140 bp
   Segment 1:  
   5697 .. 5722  =  26 bp
   9 amino acids  =  899.0 Da
Product: tryptophan oxygenase
selectable marker, required to synthesize the brown eye pigment in Drosophila
vermilion
4229 .. 5722  =  1494 bp
379 amino acids  =  44.4 kDa
6 segments  =  1140 bp
Product: tryptophan oxygenase
selectable marker, required to synthesize the brown eye pigment in Drosophila
AmpR
2408 .. 3268  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 2:  
   2408 .. 3199  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
2408 .. 3268  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 1:  signal sequence  
   3200 .. 3268  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
2408 .. 3268  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
dU6-1 promoter
7 .. 803  =  797 bp
RNA polymerase III promoter for Drosophila U6-1 snRNA (Port et al., 2014)
dU6-1 promoter
7 .. 803  =  797 bp
RNA polymerase III promoter for Drosophila U6-1 snRNA (Port et al., 2014)
ori
1649 .. 2237  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
1649 .. 2237  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
f1 ori
3399 .. 3854  =  456 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
f1 ori
3399 .. 3854  =  456 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
AmpR promoter
3269 .. 3373  =  105 bp
AmpR promoter
3269 .. 3373  =  105 bp
gRNA scaffold
822 .. 897  =  76 bp
guide RNA scaffold for the Streptococcus pyogenes CRISPR/Cas9 system
gRNA scaffold
822 .. 897  =  76 bp
guide RNA scaffold for the Streptococcus pyogenes CRISPR/Cas9 system
attB
6037 .. 6106  =  70 bp
attB site for the φC31 integrase (Groth et al., 2000)
attB
6037 .. 6106  =  70 bp
attB site for the φC31 integrase (Groth et al., 2000)
polIII terminator
898 .. 903  =  6 bp
RNA polymerase III transcription terminator
polIII terminator
898 .. 903  =  6 bp
RNA polymerase III transcription terminator
ORF:  4726 .. 4974  =  249 bp
ORF:  82 amino acids  =  9.0 kDa
ORF:  261 .. 797  =  537 bp
ORF:  178 amino acids  =  19.9 kDa
ORF:  2538 .. 2804  =  267 bp
ORF:  88 amino acids  =  9.2 kDa
ORF:  4588 .. 5181  =  594 bp
ORF:  197 amino acids  =  23.5 kDa
ORF:  5455 .. 5709  =  255 bp
ORF:  84 amino acids  =  9.7 kDa
ORF:  6340 .. 6618  =  279 bp
ORF:  92 amino acids  =  11.0 kDa
ORF:  2408 .. 3268  =  861 bp
ORF:  286 amino acids  =  31.6 kDa
ORF:  6098 .. 6421  =  324 bp
ORF:  107 amino acids  =  12.2 kDa
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