LOCUS       Exported               14976 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pGZ-DSB-CO, complete sequence.
ACCESSION   KY447298
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 14976)
  AUTHORS   Zapotoczny G, Sekelsky J.
  TITLE     Human Cell Assays for Synthesis-Dependent Strand Annealing and 
            Crossing Over During Double-Strand Break Repair
  JOURNAL   G3 (Bethesda) (2017) In press
  PUBMED    28179392
REFERENCE   2  (bases 1 to 14976)
  AUTHORS   Zapotoczny G, Sekelsky J.
  TITLE     Direct Submission
  JOURNAL   Submitted (10-JAN-2017) Department of Biology, University of North 
            Carolina, 133 N Medical Dr, Chapel Hill, NC 27599, USA
REFERENCE   3  (bases 1 to 14976)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..14976
                     /organism="Cloning vector pGZ-DSB-CO"
                     /mol_type="other DNA"
                     /db_xref="taxon:1958936"
     regulatory      116..496
                     /regulatory_class="enhancer"
                     /note="CMV immediate-early enhancer"
     enhancer        116..495
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     regulatory      497..1752
                     /regulatory_class="promoter"
                     /note="chicken beta-actin"
     promoter        498..773
                     /label=chicken beta-actin promoter
     intron          774..1782
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     misc_feature    1852..2865
                     /note="nls-eGFP with an I-SceI recognition site inserted; 
                     SceGFP"
     CDS             1867..1887
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1909..1929
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     regulatory      2143..2152
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     polyA_signal    2991..3112
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     gene            complement(3143..4891)
                     /gene="PGK1"
                     /label=PGK1
     CDS             complement(3773..4372)
                     /codon_start=1
                     /gene="PGK1"
                     /product="puromycin resistance protein"
                     /label=PGK1
                     /protein_id="AQR57379.1"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     promoter        complement(4391..4888)
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     polyA_site      5041..5096
                     /label=-beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal"
     CDS             complement(5445..6155)
                     /codon_start=1
                     /product="mCherry"
                     /label=mCherry
                     /protein_id="AQR57378.1"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     regulatory      6152..6161
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     misc_feature    complement(6164..7869)
                     /label=beta globin intron (with iGFP)
                     /note="beta globin intron (with iGFP)"
     misc_feature    complement(6393..7392)
                     /label=iGFP (lacks promoter and 5 end)
                     /note="iGFP (lacks promoter and 5 end)"
     CDS             complement(6502..7221)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     regulatory      7218..7227
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     polyA_signal    7422..7503
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     protein_bind    8052..8070
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    8078..8096
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     regulatory      complement(8096..8617)
                     /regulatory_class="promoter"
                     /note="CMV"
     enhancer        8296..8675
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     complement(8770..8786)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    8794..8810
                     /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(8818..8848)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    8863..8884
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      8989..9124
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     misc_feature    9128..10847
                     /label=mouse HPRT homology arm
                     /note="mouse HPRT homology arm"
     promoter        complement(10884..10902)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(10909..10925)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     rep_origin      11067..11522
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     promoter        11548..11652
                     /gene="bla"
                     /label=AmpR promoter
     CDS             11653..12513
                     /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"
     rep_origin      12684..13272
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     protein_bind    13560..13581
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        13596..13626
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    13634..13650
                     /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)."
     primer_bind     13658..13674
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        13695..13713
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     13750..13766
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    13785..14976
                     /label=mouse HPRT homology arm
                     /note="mouse HPRT homology arm"