LOCUS       Exported                2541 bp ds-DNA     circular SYN 13-MAY-2021
DEFINITION  gRNA cloning vector for in vitro transcription of target gRNA.
ACCESSION   .
VERSION     .
KEYWORDS    pT7-gRNA
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2541)
  AUTHORS   Jao LE, Wente SR, Chen W
  TITLE     Efficient multiplex biallelic zebrafish genome editing using a 
            CRISPR nuclease system.
  JOURNAL   Proc Natl Acad Sci U S A. 2013 Aug 5.
  PUBMED    23918387
REFERENCE   2  (bases 1 to 2541)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Proc Natl 
            Acad Sci U S A. 2013 Aug 5."
FEATURES             Location/Qualifiers
     source          1..2541
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     180..202
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     194..211
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     195..211
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_RNA        complement(239..314)
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     primer_bind     complement(347..366)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(348..366)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(390..406)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(390..406)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(403..425)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    414..430
                     /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(438..468)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    483..504
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(621..638)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(792..1380)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(872..891)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(1551..2411)
                     /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     2174..2193
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2412..2516)
                     /gene="bla"
                     /label=AmpR promoter