LOCUS       Exported                6997 bp DNA     circular SYN 24-JUL-2020
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6997)
  AUTHORS   MIT
  TITLE     Direct Submission
  JOURNAL   Exported Jul 21, 2021 from SnapGene 5.3.2
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6997
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     protein_bind    820..838
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    841..859
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     CDS             996..1019
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             1020..2909
                     /codon_start=1
                     /label=ePAB-B
                     /note="ePAB-B"
                     /translation="MNATRAEYPLASLYIGDLHPDVTEAMLYEKFSPAGPIMSIRVCRD
                     IATRRSLGYAYINFQQPADAERALDTMNFEVIKGRPIRIMWSQRDPGLRKSGVGNVFIK
                     NLDDSIDNKALYDTFSAFGDILSCKVVCDEYGSRGYGFVHFETQEAANRAIQTMNGMLL
                     NDRKVFVGHFKSRRERELEYGAKVMEFTNVYIKNFGEDMDDKRLKEIFSAFGNTLSVKV
                     MMDNSGRSRGFGFVNYGNHEEAQKAVTEMNGKEVNGRMVYVGRAQKRIERQGELKRKFE
                     QIKQERINRYQGVNLYVKNLDDGIDDDRLRKEFSPYGTITSTKVMTEGGHSKGFGFVCF
                     SSPEEATKAVTEMNGRIVSTKPLYVALAQRKEERKAILTNQYMQRLATMRAMPGPLLGS
                     FQQPANYFLPTMPQPSNRAFYSPNPVAPVRPAPQWASHQSRPPQYQPPAPLMRAVPPRR
                     MHSNISTMKQASTQVPRVPLQSQRVANIGTQTAGARAQVNASIMRAMPHYKYSCGVRNV
                     QPIGSSAHLQQVLEPAVLMQGQEPLTASLLAAAPLQEQKQILGERIYPLIHEMHPTLAG
                     KITGMLLEIDNSELLHMLESPESLHSKVEEAVAVLQAHQAKESAPKSAPQSLI"
     misc_feature    2910..2929
                     /label=3' end preferred cloning region
     polyA_signal    2955..3179
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     protein_bind    3463..3510
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     CDS             3518..4540
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     polyA_signal    4670..4791
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4840..4856)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    4864..4880
                     /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(4888..4918)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4933..4954
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(5242..5830)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(6001..6861)
                     /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"
     promoter        complement(6862..6966)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc
      841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga
      901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga
      961 ctctagcgtt taaacttaag cttgccgcca ccatggatta caaggatgac gacgataaga
     1021 tgaacgcaac cagagccgaa tacccgctgg catctcttta tataggggac ctccaccccg
     1081 acgtgacgga agcgatgctg tatgagaagt tttctccagc ggggcccatc atgtctataa
     1141 gagtgtgtcg agatattgct actcgccggt cccttggcta tgcctacatt aatttccagc
     1201 agccggctga tgctgagcgg gccttggata caatgaattt tgaggttatt aaaggcaggc
     1261 caatccgaat catgtggtcc cagagagatc cgggccttcg aaaatcgggt gtagggaatg
     1321 tatttattaa aaatttagac gactccatcg acaacaaagc cctgtatgac actttctcag
     1381 cgtttggaga tattttatct tgcaaagttg tgtgcgatga atacggatct cgaggttatg
     1441 gtttcgtaca ttttgagacc caagaagctg ccaatcgagc gattcagact atgaatggga
     1501 tgttgctcaa cgaccgcaaa gtgtttgtgg gccattttaa atcacggagg gaaagagaat
     1561 tagaatatgg ggcaaaagtc atggaattta ccaacgtgta catcaagaac ttcggggagg
     1621 atatggacga taaaagactt aaagaaatat tttctgcctt tggaaatacg ttaagcgtta
     1681 aagtaatgat ggataatagt ggtcgatccc ggggttttgg ttttgttaat tatggaaatc
     1741 acgaagaagc acaaaaagca gttaccgaaa tgaatggtaa agaagtaaac ggtcgaatgg
     1801 tttacgtagg cagagctcag aagaggatag aacggcaggg cgagttaaaa cgcaagttcg
     1861 aacaaatcaa gcaagaacga atcaaccgct atcagggtgt taatttgtat gttaaaaacc
     1921 tggacgatgg tatcgatgat gaccggctca gaaaggagtt ctcgccgtat ggtaccatta
     1981 ccagtacaaa ggtaatgact gaaggaggcc atagtaaagg ctttggtttt gtttgcttct
     2041 cctccccaga agaagccact aaagcagtca cagaaatgaa cgggaggata gtcagtacga
     2101 agcctctata tgttgcactt gcacaaagga aggaagaacg gaaggcaatt ctaaccaatc
     2161 agtacatgca gaggctagcc actatgaggg ccatgcctgg tcctctcctt gggtccttcc
     2221 agcaacctgc taattatttc ttgcctacaa tgccacagcc ttcaaacaga gcattttata
     2281 gtccaaatcc tgttgctcct gtccgtccag ctcctcagtg ggcttcccac cagtccaggc
     2341 ctccacagta tcaaccacca gccccattaa tgagggcggt gccaccaaga aggatgcatt
     2401 caaatatcag tacaatgaag caggcatcaa cacaagtacc cagggtgcct ctacagagtc
     2461 aaagagtagc aaacattggc actcaaacag ctggagctcg ggcccaagta aacgcgtcca
     2521 taatgcgcgc aatgccacat tacaagtatt cctgtggcgt gaggaacgtg cagcctatag
     2581 gaagctctgc tcatctgcaa caggtactgg aacctgcagt tctgatgcaa ggacaagaac
     2641 cgctaactgc atctctgctt gcagctgccc cactgcagga acagaaacaa atactcgggg
     2701 aacgtattta cccgttaata catgaaatgc atccgacgtt agccggaaag ataacgggaa
     2761 tgctgctgga aattgataac tctgagctac tgcatatgct ggagtcacca gagtctctgc
     2821 attcaaaggt ggaagaggca gtggcggtcc tgcaagccca ccaagccaag gaaagtgctc
     2881 cgaaaagtgc tccgcagtct ctgatctgac gctcgagtct agagggcccg tttaaacccg
     2941 ctgatcagcc tcgactgtgc cttctagttg ccagccatct gttgtttgcc cctcccccgt
     3001 gccttccttg accctggaag gtgccactcc cactgtcctt tcctaataaa atgaggaaat
     3061 tgcatcgcat tgtctgagta ggtgtcattc tattctgggg ggtggggtgg ggcaggacag
     3121 caagggggag gattgggaag acaatagcag gcatgctggg gatgcggtgg gctctatggc
     3181 ttctgaggcg gaaagaacca gctggggctc tagggggtat ccccacgcgc cctgtagcgg
     3241 cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc
     3301 cctagcgccc gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc
     3361 ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct
     3421 cgaccccaaa aaacttgatt agggtgatgg ttcacgtacc tagaagttcc tattccgaag
     3481 ttcctattct ctagaaagta taggaacttc cttggccaaa aagcctgaac tcaccgcgac
     3541 gtctgtcgag aagtttctga tcgaaaagtt cgacagcgtc tccgacctga tgcagctctc
     3601 ggagggcgaa gaatctcgtg ctttcagctt cgatgtagga gggcgtggat atgtcctgcg
     3661 ggtaaatagc tgcgccgatg gtttctacaa agatcgttat gtttatcggc actttgcatc
     3721 ggccgcgctc ccgattccgg aagtgcttga cattggggaa ttcagcgaga gcctgaccta
     3781 ttgcatctcc cgccgtgcac agggtgtcac gttgcaagac ctgcctgaaa ccgaactgcc
     3841 cgctgttctg cagccggtcg cggaggccat ggatgcgatc gctgcggccg atcttagcca
     3901 gacgagcggg ttcggcccat tcggaccgca aggaatcggt caatacacta catggcgtga
     3961 tttcatatgc gcgattgctg atccccatgt gtatcactgg caaactgtga tggacgacac
     4021 cgtcagtgcg tccgtcgcgc aggctctcga tgagctgatg ctttgggccg aggactgccc
     4081 cgaagtccgg cacctcgtgc acgcggattt cggctccaac aatgtcctga cggacaatgg
     4141 ccgcataaca gcggtcattg actggagcga ggcgatgttc ggggattccc aatacgaggt
     4201 cgccaacatc ttcttctgga ggccgtggtt ggcttgtatg gagcagcaga cgcgctactt
     4261 cgagcggagg catccggagc ttgcaggatc gccgcggctc cgggcgtata tgctccgcat
     4321 tggtcttgac caactctatc agagcttggt tgacggcaat ttcgatgatg cagcttgggc
     4381 gcagggtcga tgcgacgcaa tcgtccgatc cggagccggg actgtcgggc gtacacaaat
     4441 cgcccgcaga agcgcggccg tctggaccga tggctgtgta gaagtactcg ccgatagtgg
     4501 aaaccgacgc cccagcactc gtccgagggc aaaggaatag cacgtactac gagatttcga
     4561 ttccaccgcc gccttctatg aaaggttggg cttcggaatc gttttccggg acgccggctg
     4621 gatgatcctc cagcgcgggg atctcatgct ggagttcttc gcccacccca acttgtttat
     4681 tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt
     4741 tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt atcatgtctg
     4801 tataccgtcg acctctagct agagcttggc gtaatcatgg tcatagctgt ttcctgtgtg
     4861 aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc
     4921 ctggggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt
     4981 ccagtcggga aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg
     5041 cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt
     5101 tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc
     5161 aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa
     5221 aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa
     5281 tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc
     5341 ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc
     5401 cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag
     5461 ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga
     5521 ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc
     5581 gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac
     5641 agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg
     5701 cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca
     5761 aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa
     5821 aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa
     5881 ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt
     5941 aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag
     6001 ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat
     6061 agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc
     6121 cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa
     6181 ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca
     6241 gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa
     6301 cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt
     6361 cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc
     6421 ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact
     6481 catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc
     6541 tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg
     6601 ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct
     6661 catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc
     6721 cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag
     6781 cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac
     6841 acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg
     6901 ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt
     6961 tccgcgcaca tttccccgaa aagtgccacc tgacgtc
//
