Resources
Plasmid Files

sgRNA(MS2) cloning backbone

Zhang lab vector for expressing a single guide RNA (sgRNA) containing two MS2 stem loops.

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sgRNA(MS2) cloning backbone Sequence and MapsgRNA(MS2) cloning backbone.dna
Map and Sequence File   
Sequence Author:  Zhang Lab / Addgene #61424
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 BbsI (251) BbsI (273) sgRNA(MS2) cloning backbone 3008 bp
BbsI  (251)
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  (273)
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.
AmpR
1333 .. 2193  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 1:  signal sequence  
   1333 .. 1401  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
1333 .. 2193  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 2:  
   1402 .. 2193  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
1333 .. 2193  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
ori
2364 .. 2952  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
2364 .. 2952  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
f1 ori
491 .. 946  =  456 bp
f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis
f1 ori
491 .. 946  =  456 bp
f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis
U6 promoter
7 .. 247  =  241 bp
RNA polymerase III promoter for human U6 snRNA
U6 promoter
7 .. 247  =  241 bp
RNA polymerase III promoter for human U6 snRNA
gRNA scaffold
274 .. 409  =  136 bp
guide RNA scaffold for the Streptococcus pyogenes
CRISPR/Cas9 system, modified by the insertion of
two MS2 stem loops
gRNA scaffold
274 .. 409  =  136 bp
guide RNA scaffold for the Streptococcus pyogenes
CRISPR/Cas9 system, modified by the insertion of
two MS2 stem loops
AmpR promoter
1228 .. 1332  =  105 bp
AmpR promoter
1228 .. 1332  =  105 bp
polIII terminator
410 .. 415  =  6 bp
RNA polymerase III transcription terminator
polIII terminator
410 .. 415  =  6 bp
RNA polymerase III transcription terminator
MS2 stem loop
291 .. 309  =  19 bp
stem loop that binds the bacteriophage MS2 coat
protein
MS2 stem loop
291 .. 309  =  19 bp
stem loop that binds the bacteriophage MS2 coat
protein
MS2 stem loop
361 .. 379  =  19 bp
stem loop that binds the bacteriophage MS2 coat
protein
MS2 stem loop
361 .. 379  =  19 bp
stem loop that binds the bacteriophage MS2 coat
protein
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