LOCUS       Exported                2962 bp ds-DNA     circular SYN 13-MAY-2021
DEFINITION  Backbone for cloning guides that are compatible with LwaCas13a. 
            Contains a 5 direct repeat. Clone using BbsI (BpiI). F overhang 
            AAAC. R overhang AAAA.
ACCESSION   .
VERSION     .
KEYWORDS    pC0040-LwaCas13a crRNA backbone
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2962)
  AUTHORS   Cox DBT, Gootenberg JS, Abudayyeh OO, Franklin B, Kellner MJ, Joung 
            J, Zhang F
  TITLE     RNA editing with CRISPR-Cas13.
  JOURNAL   Science. 2017 Oct 25. pii: eaaq0180. doi: 10.1126/science.aaq0180.
  PUBMED    29070703
REFERENCE   2  (bases 1 to 2962)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Science. 
            2017 Oct 25. pii: eaaq0180. doi: 10.1126/science.aaq0180."
FEATURES             Location/Qualifiers
     source          1..2962
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     rep_origin      complement(38..626)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(118..137)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(797..1657)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     1420..1439
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1658..1762)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     1830..1848
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(1886..1908)
                     /label=pGEX 3
                     /note="pGEX vectors, reverse primer"
     primer_bind     2008..2027
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2221..2243
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2235..2252
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2236..2252
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        complement(2344..2584)
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA"
     primer_bind     complement(2394..2413)
                     /label=LKO.1 5
                     /note="Human U6 promoter, forward primer"
     primer_bind     complement(2564..2584)
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     complement(2598..2614)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2598..2614)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2611..2633)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2622..2638
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(2646..2676)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2691..2712
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2829..2846)
                     /label=L4440
                     /note="L4440 vector, forward primer"