LOCUS       sequence-66850-e        5044 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid for cloning and constitutive or doxycycline-inducible 
            expression of cDNAs. Confers resistance to puromycin. Use T-REx 
            cells for doxycycline-inducible expression..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5044)
  AUTHORS   Britton S, Coates J, Jackson SP
  TITLE     A new method for high-resolution imaging of Ku foci to decipher 
            mechanisms of DNA double-strand break repair.
  JOURNAL   J Cell Biol. 2013 Jul 29.
  PUBMED    23897892
REFERENCE   2  (bases 1 to 5044)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "J Cell 
            Biol. 2013 Jul 29."
COMMENT     Sequence Label: pICE
FEATURES             Location/Qualifiers
     source          1..5044
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      7..16
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(125..142)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    131..355
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      401..829
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(488..507)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     698..719
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(838..858)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        843..1172
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      1023..1158
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     1085..1104
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             1224..1823
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(1224..1243)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     1720..1740
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    1953..2074
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1990..2009)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2044..2063
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(2124..2140)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2124..2140)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2137..2159)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2148..2164
                     /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(join(2172..2178,2179..2196,2197..2202))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2217..2238
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2355..2372)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2526..3111)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2606..2625)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3282..4073,4074..4142))
                     /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     3905..3924
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4143..4247)
                     /gene="bla"
                     /label=AmpR promoter
     enhancer        4319..4698
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        4699..4902
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     4853..4873
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    4904..4922
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    4925..4943
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     4953..4977
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
ORIGIN
        1 aagcttgcca ccatggtgct gcacgagtac gtgaacgccg ccggcatcac agacaaatgc
       61 ggccgcgaat tcggatccgg taccctcgag acgcgtctgc agtctagata aacccgctga
      121 tcagcctcga ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct
      181 tccttgaccc tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca
      241 tcgcattgtc tgagtaggtg tcattctatt ctggggggtg gggtggggca ggacagcaag
      301 ggggaggatt gggaagacaa tagcaggcat gctggggatg cggtgggctc tatggcttct
      361 gaggcggaaa gaaccagctg gggctctagg gggtatcccc acgcgccctg tagcggcgca
      421 ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta
      481 gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt
      541 caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac
      601 cccaaaaaac ttgattaggg tgatggttca cgtagtgggc catcgccctg atagacggtt
      661 tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga
      721 acaacactca accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg
      781 gcctattggt taaaaaatga gctgatttaa caaaaattta acgcgaatta attctgtgga
      841 atgtgtgtca gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa
      901 gcatgcatct caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca
      961 gaagtatgca aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc
     1021 ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt
     1081 tttttattta tgcagaggcc gaggccgcct ctgcctctga gctattccag aagtagtgag
     1141 gaggcttttt tggaggccta ggcttttgca aaaagctccc gggagcttgt atatccattt
     1201 tcggatctga tcagcacgtg accatgaccg agtacaagcc cacggtgcgc ctcgccaccc
     1261 gcgacgacgt ccccagggcc gtacgcaccc tcgccgccgc gttcgccgac taccccgcca
     1321 cgcgccacac cgtcgatccg gaccgccaca tcgagcgggt caccgagctg caagaactct
     1381 tcctcacgcg cgtcgggctc gacatcggca aggtgtgggt cgcggacgac ggcgccgcgg
     1441 tggcggtctg gaccacgccg gagagcgtcg aagcgggggc ggtgttcgcc gagatcggcc
     1501 cgcgcatggc cgagttgagc ggttcccggc tggccgcgca gcaacagatg gaaggcctcc
     1561 tggcgccgca ccggcccaag gagcccgcgt ggttcctggc caccgtcggc gtctcgcccg
     1621 accaccaggg caagggtctg ggcagcgccg tcgtgctccc cggagtggag gcggccgagc
     1681 gcgccggggt gcccgccttc ctggagacct ccgcgccccg caacctcccc ttctacgagc
     1741 ggctcggctt caccgtcacc gccgacgtcg aggtgcccga aggaccgcgc acctggtgca
     1801 tgacccgcaa gcccggtgcc tgacacgtgc tacgagattt cgattccacc gccgccttct
     1861 atgaaaggtt gggcttcgga atcgttttcc gggacgccgg ctggatgatc ctccagcgcg
     1921 gggatctcat gctggagttc ttcgcccacc ccaacttgtt tattgcagct tataatggtt
     1981 acaaataaag caatagcatc acaaatttca caaataaagc atttttttca ctgcattcta
     2041 gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctggctagcc gtcgacctct
     2101 agctagagct tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc
     2161 acaattccac acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga
     2221 gtgagctaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg
     2281 tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg
     2341 cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg
     2401 gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga
     2461 aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg
     2521 gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag
     2581 aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc
     2641 gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg
     2701 ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt
     2761 cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc
     2821 ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc
     2881 actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg
     2941 tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca
     3001 gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc
     3061 ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct
     3121 ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg
     3181 gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt
     3241 aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt
     3301 gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc
     3361 gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg
     3421 cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc
     3481 gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg
     3541 gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca
     3601 ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga
     3661 tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct
     3721 ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg
     3781 cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca
     3841 accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata
     3901 cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct
     3961 tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact
     4021 cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa
     4081 acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc
     4141 atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga
     4201 tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga
     4261 aaagtgccac ctgacgtcga cggatcggga gatcctgtac gggccagata tccgcgttga
     4321 cattgattat tgactagtta ttaatagtaa tcaattacgg ggtcattagt tcatagccca
     4381 tatatggagt tccgcgttac ataacttacg gtaaatggcc cgcctggctg accgcccaac
     4441 gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc aatagggact
     4501 ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc agtacatcaa
     4561 gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg gcccgcctgg
     4621 cattatgccc agtacatgac cttatgggac tttcctactt ggcagtacat ctacgtatta
     4681 gtcatcgcta ttaccatggt gatgcggttt tggcagtaca tcaatgggcg tggatagcgg
     4741 tttgactcac ggggatttcc aagtctccac cccattgacg tcaatgggag tttgttttgg
     4801 caccaaaatc aacgggactt tccaaaatgt cgtaacaact ccgccccatt gacgcaaatg
     4861 ggcggtaggc gtgtacggtg ggaggtctat ataagcagag ctctccctat cagtgataga
     4921 gatctcccta tcagtgatag agatcgtcga cgagctcgtt tagtgaaccg tcagatcgcc
     4981 tggagacgcc atccacgctg ttttgacctc catagaagac accgggaccg atccagcctc
     5041 cggc
//
