LOCUS       Exported                9050 bp ds-DNA     circular SYN 26-FEB-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  recombinant plasmid
REFERENCE   1  (bases 1 to 9050)
  AUTHORS   Simon
  TITLE     Direct Submission
  JOURNAL   Exported Wednesday, Feb 26, 2025 from SnapGene 5.0.8
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..9050
                     /organism="recombinant plasmid"
                     /mol_type="other DNA"
     promoter        complement(1..105)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(131..586)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     728..744
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    1296..1320
                     /label=SEQ_CMV_FW
     enhancer        1360..1724
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1725..1928
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     protein_bind    1930..1948
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1951..1969
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     misc_feature    2010..2036
                     /label=SEQ_FW_PBtetr
     CDS             2091..2288
                     /codon_start=1
                     /label=TTHA1718
                     /translation="MLKLKVEGMTCNHCVMAVTKALKKVPGVEKVEVSLEKGEALVEGT
                     ADPKALVQAVEEEGYKAEVLA"
     CDS             2289..2312
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     misc_feature    2325..2911
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     misc_feature    2924..3682
                     /label=copGFP
     promoter        3731..3942
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     LTR             3955..4223
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from human 
                     T-cell leukemia virus (HTLV) type 1"
     misc_feature    4182..4206
                     /label=SEQ_TETr_FW
     CDS             4260..4877
                     /codon_start=1
                     /gene="tetR from transposon Tn10"
                     /product="tetracycline repressor TetR"
                     /label=TetR
                     /note="TetR binds to the tetracycline operator tetO to 
                     inhibit transcription. This inhibition can be relieved by 
                     adding tetracycline or doxycycline."
                     /translation="MSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESG"
     CDS             4914..4967
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             4974..5573
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     misc_feature    complement(5017..5041)
                     /label=SEQ_TETr_RV
     polyA_signal    5620..5741
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7029..7045)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    7053..7069
                     /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(7077..7107)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7122..7143
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(7431..8019)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(8190..9050)
                     /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"
ORIGIN
        1 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata
       61 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa
      121 agtgccacct aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa
      181 atcagctcat tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa
      241 tagaccgaga tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac
      301 gtggactcca acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa
      361 ccatcaccct aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct
      421 aaagggagcc cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa
      481 gggaagaaag cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc
      541 gtaaccacca cacccgccgc gcttaatgcg ccgctacagg gcgcgtccca ttcgccattc
      601 aggctgcgca actgttggga agggcgatcg gtgcgggcct cttcgctatt acgccagctg
      661 gcgaaagggg gatgtgctgc aaggcgatta agttgggtaa cgccagggtt ttcccagtca
      721 cgacgttgta aaacgacggc cagtgagcgc gcctcgttca ttcacgtttt tgaacccgtg
      781 gaggacgggc agactcgcgg tgcaaatgtg ttttacagcg tgatggagca gatgaagatg
      841 ctcgacacgc tgcagaacac gcagctagat taaccctaga aagataatca tattgtgacg
      901 tacgttaaag ataatcatgt gtaaaattga cgcatgtgtt ttatcggtct gtatatcgag
      961 gtttatttat taatttgaat agatattaag ttttattata tttacactta catactaata
     1021 ataaattcaa caaacaattt atttatgttt atttatttat taaaaaaaac aaaaactcaa
     1081 aatttcttct ataaagtaac aaaactttta tgagggacag ccccccccca aagcccccag
     1141 ggatgtaatt acgtccctcc cccgctaggg ggcagcagcg agccgcccgg ggctccgctc
     1201 cggtccggcg ctccccccgc atccccgagc cggcagcgtg cggggacagc ccgggcacgg
     1261 ggaaggtggc acgggatcgc tttcctctga acgcttctcg ctgctctttg agcctgcaga
     1321 cacctggggg gatacgggga aaaggcctcc acggccagac tagttattaa tagtaatcaa
     1381 ttacggggtc attagttcat agcccatata tggagttccg cgttacataa cttacggtaa
     1441 atggcccgcc tggctgaccg cccaacgacc cccgcccatt gacgtcaata atgacgtatg
     1501 ttcccatagt aacgccaata gggactttcc attgacgtca atgggtggag tatttacggt
     1561 aaactgccca cttggcagta catcaagtgt atcatatgcc aagtacgccc cctattgacg
     1621 tcaatgacgg taaatggccc gcctggcatt atgcccagta catgacctta tgggactttc
     1681 ctacttggca gtacatctac gtattagtca tcgctattac catggtgatg cggttttggc
     1741 agtacatcaa tgggcgtgga tagcggtttg actcacgggg atttccaagt ctccacccca
     1801 ttgacgtcaa tgggagtttg ttttggaacc aaaatcaacg ggactttcca aaatgtcgta
     1861 acaactccgc cccattgacg caaatgggcg gtaggcgtgt acggtgggag gtctatataa
     1921 gcagagctct ccctatcagt gatagagatc tccctatcag tgatagagat cgtcgacgag
     1981 ctcgtttagt gaaccgtcag atcgcctgga gacgccatcc acgctgtttt gacctccata
     2041 gaagacaccg ggaccgatcc agcctccgga ctctagcggt ctaggctagc atgctgaagc
     2101 tcaaggtgga aggcatgacc tgcaaccact gcgtgatggc ggtgaccaag gccctgaaga
     2161 aggtccccgg cgtggagaag gtggaggtct ccctagaaaa gggggaggcc ctggtggagg
     2221 ggacggccga ccccaaggcc ctcgtccagg cggtggagga ggaagggtac aaggccgagg
     2281 ttctggccga ctacaaggac gacgacgaca agtgagcggc cgcgcccctc tccctccccc
     2341 ccccctaacg ttactggccg aagccgcttg gaataaggcc ggtgtgcgtt tgtctatatg
     2401 ttattttcca ccatattgcc gtcttttggc aatgtgaggg cccggaaacc tggccctgtc
     2461 ttcttgacga gcattcctag gggtctttcc cctctcgcca aaggaatgca aggtctgttg
     2521 aatgtcgtga aggaagcagt tcctctggaa gcttcttgaa gacaaacaac gtctgtagcg
     2581 accctttgca ggcagcggaa ccccccacct ggcgacaggt gcctctgcgg ccaaaagcca
     2641 cgtgtataag atacacctgc aaaggcggca caaccccagt gccacgttgt gagttggata
     2701 gttgtggaaa gagtcaaatg gctctcctca agcgtattca acaaggggct gaaggatgcc
     2761 cagaaggtac cccattgtat gggatctgat ctggggcctc ggtgcacatg ctttacatgt
     2821 gtttagtcga ggttaaaaaa acgtctaggc cccccgaacc acggggacgt ggttttcctt
     2881 tgaaaaacac gatgataata tggccacaac catggcgtcc ggaatggaga gcgacgagag
     2941 cggcctgccc gccatggaga tcgagtgccg catcaccggc accctgaacg gcgtggagtt
     3001 cgagctggtg ggcggcggag agggcacccc caagcagggc cgcatgacca acaagatgaa
     3061 gagcaccaaa ggcgccctga ccttcagccc ctacctgctg agccacgtga tgggctacgg
     3121 cttctaccac ttcggcacct accccagcgg ctacgagaac cccttcctgc acgccatcaa
     3181 caacggcggc tacaccaaca cccgcatcga gaagtacgag gacggcggcg tgctgcacgt
     3241 gagcttcagc taccgctacg aggccggccg cgtgatcggc gacttcaagg tggtgggcac
     3301 cggcttcccc gaggacagcg tgatcttcac cgacaagatc atccgcagca acgccaccgt
     3361 ggagcacctg caccccatgg gcgataacgt gctggtgggc agcttcgccc gcaccttcag
     3421 cctgcgcgac ggcggctact acagcttcgt ggtggacagc cacatgcact tcaagagcgc
     3481 catccacccc agcatcctgc agaacggggg ccccatgttc gccttccgcc gcgtggagga
     3541 gctgcacagc aacaccgagc tgggcatcgt ggagtaccag cacgccttca agacccccat
     3601 cgccttcgcc agatcccgcg ctcagtcgtc caattctgcc gtggacggca ccgccggacc
     3661 cggctccacc ggatctcgct agagctgaat ctaagtcgac ggatctgcga tcgctccggt
     3721 gcccgtcagt gggcagagcg cacatcgccc acagtccccg agaagttggg gggaggggtc
     3781 ggcaattgaa cgggtgccta gagaaggtgg cgcggggtaa actgggaaag tgatgtcgtg
     3841 tactggctcc gcctttttcc cgagggtggg ggagaaccgt atataagtgc agtagtcgcc
     3901 gtgaacgttc tttttcgcaa cgggtttgcc gccagaacac agctgaagct tcgaggggct
     3961 cgcatctctc cttcacgcgc ccgccgccct acctgaggcc gccatccacg ccggttgagt
     4021 cgcgttctgc cgcctcccgc ctgtggtgcc tcctgaactg cgtccgccgt ctaggtaagt
     4081 ttaaagctca ggtcgagacc gggcctttgt ccggcgctcc cttggagcct acctagactc
     4141 agccggctct ccacgctttg cctgaccctg cttgctcaac tctacgtctt tgtttcgttt
     4201 tctgttctgc gccgttacag atccaagctg tgaccggcgc ctactctaga gccgccacca
     4261 tgtccagatt agataaaagt aaagtgatta acagcgcatt agagctgctt aatgaggtcg
     4321 gaatcgaagg tttaacaacc cgtaaactcg cccagaagct aggtgtagag cagcctacat
     4381 tgtattggca tgtaaaaaat aagcgggctt tgctcgacgc cttagccatt gagatgttag
     4441 ataggcacca tactcacttt tgccctttag aaggggaaag ctggcaagat tttttacgta
     4501 ataacgctaa aagttttaga tgtgctttac taagtcatcg cgatggagca aaagtacatt
     4561 taggtacacg gcctacagaa aaacagtatg aaactctcga aaatcaatta gcctttttat
     4621 gccaacaagg tttttcacta gagaatgcat tatatgcact cagcgctgtg gggcatttta
     4681 ctttaggttg cgtattggaa gatcaagagc atcaagtcgc taaagaagaa agggaaacac
     4741 ctactactga tagtatgccg ccattattac gacaagctat cgaattattt gatcaccaag
     4801 gtgcagagcc agccttctta ttcggccttg aattgatcat atgcggatta gaaaaacaac
     4861 ttaaatgtga aagtgggtcc gcgtacagcg gatcccggga gttcagatct tatgagggca
     4921 gaggaagtct tctaacatgc ggtgacgtgg aggagaatcc cggccctgaa ttcatgaccg
     4981 agtacaagcc cacggtgcgc ctcgccaccc gcgacgacgt ccccagggcc gtacgcaccc
     5041 tcgccgccgc gttcgccgac taccccgcca cgcgccacac cgtcgatccg gaccgccaca
     5101 tcgagcgggt caccgagctg caagaactct tcctcacgcg cgtcgggctc gacatcggca
     5161 aggtgtgggt cgcggacgac ggcgccgcgg tggcggtctg gaccacgccg gagagcgtcg
     5221 aagcgggggc ggtgttcgcc gagatcggcc cgcgcatggc cgagttgagc ggttcccggc
     5281 tggccgcgca gcaacagatg gaaggcctcc tggcgccgca ccggcccaag gagcccgcgt
     5341 ggttcctggc caccgtcggc gtctcgcccg accaccaggg caagggtctg ggcagcgccg
     5401 tcgtgctccc cggagtggag gcggccgagc gcgccggggt gcccgccttc ctggagacct
     5461 ccgcgccccg caacctcccc ttctacgagc ggctcggctt caccgtcacc gccgacgtcg
     5521 aggtgcccga aggaccgcgc acctggtgca tgacccgcaa gcccggtgcc tgagtcgaca
     5581 atcaacctct ggattacaaa atttgtggtt aactaactaa acttgtttat tgcagcttat
     5641 aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg
     5701 cattctagtt gtggtttgtc caaactcatc aatgtatctt atcatgtctg gaattgactc
     5761 aaatgatgtc aattagtcta tcagaagctc atctggtctc ccttccgggg gacaagacat
     5821 ccctgtttaa tatttaaaca gcagtgttcc caaactgggt tcttatatcc cttgctctgg
     5881 tcaaccaggt tgcagggttt cctgtcctca caggaacgaa gtccctaaag aaacagtggc
     5941 agccaggttt agccccggaa ttgactggat tcctttttta gggcccattg gtatggcttt
     6001 ttccccgtat ccccccaggt gtctgcaggc tcaaagagca gcgagaagcg ttcagaggaa
     6061 agcgatcccg tgccaccttc cccgtgcccg ggctgtcccc gcacgctgcc ggctcgggga
     6121 tgcgggggga gcgccggacc ggagcggagc cccgggcggc tcgctgctgc cccctagcgg
     6181 gggagggacg taattacatc cctgggggct ttgggggggg gctgtccctg atatctataa
     6241 caagaaaata tatatataat aagttatcac gtaagtagaa catgaaataa caatataatt
     6301 atcgtatgag ttaaatctta aaagtcacgt aaaagataat catgcgtcat tttgactcac
     6361 gcggtcgtta tagttcaaaa tcagtgacac ttaccgcatt gacaagcacg cctcacggga
     6421 gctccaagcg gcgactgaga tgtcctaaat gcacagcgac ggattcgcgc tatttagaaa
     6481 gagagagcaa tatttcaaga atgcatgcgt caattttacg cagactatct ttctagggtt
     6541 aatctagctg catcaggatc atatcgtcgg gtcttttttc cggctcagtc atcgcccaag
     6601 ctggcgctat ctgggcatcg gggaggaaga agcccgtgcc ttttcccgcg aggttgaagc
     6661 ggcatggaaa gagtttgccg aggatgactg ctgctgcatt gacgttgagc gaaaacgcac
     6721 gtttaccatg atgattcggg aaggtgtggc catgcacgcc tttaacggtg aactgttcgt
     6781 tcaggccacc tgggatacca gttcgtcgcg gcttttccgg acacagttcc ggatggtcag
     6841 cccgaagcgc atcagcaacc cgaacaatac cggcgacagc cggaactgcc gtgccggtgt
     6901 gcagattaat gacagcggtg cggcgctggg atattacgtc agcgaggacg ggtatcctgg
     6961 ctggatgccg cagaaatgga catggatacc ccgtgagtta cccggcgggc gcgcttggcg
     7021 taatcatggt catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac
     7081 atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca
     7141 ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat
     7201 taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc
     7261 tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca
     7321 aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca
     7381 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     7441 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     7501 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     7561 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     7621 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     7681 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     7741 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     7801 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     7861 tacactagaa ggacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     7921 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
     7981 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     8041 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
     8101 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
     8161 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
     8221 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact
     8281 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc
     8341 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt
     8401 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta
     8461 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg
     8521 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt
     8581 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc
     8641 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt
     8701 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc
     8761 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc
     8821 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa
     8881 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac
     8941 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa
     9001 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat
//
