LOCUS       Exported                8287 bp ds-DNA     circular SYN 19-JAN-2021
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  recombinant plasmid
REFERENCE   1  (bases 1 to 8287)
  AUTHORS   wallv
  TITLE     Direct Submission
  JOURNAL   Exported Tuesday, Jan 19, 2021 from SnapGene Viewer 5.2.4
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..8287
                     /organism="recombinant plasmid"
                     /mol_type="other DNA"
     rep_origin      204..1993
                     /label=oriP
                     /note="Epstein-Barr virus oriP replication origin (Yates et
                     al., 2000)"
     enhancer        2231..2534
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2535..2738
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     protein_bind    2804..2828
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             2831..3754
                     /codon_start=1
                     /label=TPA-CoV-2-S(318-529)-3C-His8-SBP
                     /translation="MDAMKRGLCCVLLLCGAVFVSPSASFRVQPTESIVRFPNITNLCP
                     FGEVFNATRFASVYAWNRKRISNCVADYSVLYNSASFSTFKCYGVSPTKLNDLCFTNVY
                     ADSFVIRGDEVRQIAPGQTGKIADYNYKLPDDFTGCVIAWNSNNLDSKVGGNYNYLYRL
                     FRKSNLKPFERDISTEIYQAGSTPCNGVEGFNCYFPLQSYGFQPTNGVGYQPYRVVVLS
                     FELLHAPATVCGPKKGAGSSLEVLFQGPGSGSSHHHHHHHHGGSGSSMDEKTTGWRGGH
                     VVEGLAGELEQLRARLEHHPQGQREP"
     sig_peptide     2831..2905
                     /label=TPA leader
     CDS             2906..3541
                     /codon_start=1
                     /label=SARS-CoV-2 RBD(318-529)
                     /translation="FRVQPTESIVRFPNITNLCPFGEVFNATRFASVYAWNRKRISNCV
                     ADYSVLYNSASFSTFKCYGVSPTKLNDLCFTNVYADSFVIRGDEVRQIAPGQTGKIADY
                     NYKLPDDFTGCVIAWNSNNLDSKVGGNYNYLYRLFRKSNLKPFERDISTEIYQAGSTPC
                     NGVEGFNCYFPLQSYGFQPTNGVGYQPYRVVVLSFELLHAPATVCGPKK"
     CDS             3557..3580
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             3596..3619
                     /codon_start=1
                     /product="8xHis affinity tag"
                     /label=8xHis
                     /translation="HHHHHHHH"
     CDS             3638..3751
                     /codon_start=1
                     /product="streptavidin-binding peptide"
                     /label=SBP
                     /note="selected from a peptide library; binds streptavidin 
                     with nanomolar affinity (Keefe et al., 2001)"
                     /translation="MDEKTTGWRGGHVVEGLAGELEQLRARLEHHPQGQREP"
     protein_bind    complement(3753..3777)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     promoter        complement(3839..3857)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     polyA_signal    3883..4107
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      4153..4581
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     promoter        4595..4925
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4776..4911
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4992..5786
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     polyA_signal    5960..6081
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6130..6146)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    6154..6170
                     /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(6178..6208)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6223..6244
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(6532..7120)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(7291..8151)
                     /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(8152..8256)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatccgcc tttatgtgta actcttggct gaagctctta caccaatgct
       61 gggggacatg tacctcccag gggcccagga agactacggg aggctacacc aacgtcaatc
      121 agaggggcct gtgtagctac cgataagcgg accctcaaga gggcattagc aatagtgttt
      181 ataaggcccc cttgttaacc ctaaacgggt agcatatgct tcccgggtag tagtatatac
      241 tatccagact aaccctaatt caatagcata tgttacccaa cgggaagcat atgctatcga
      301 attagggtta gtaaaagggt cctaaggaac agcgatatct cccaccccat gagctgtcac
      361 ggttttattt acatggggtc aggattccac gagggtagtg aaccatttta gtcacaaggg
      421 cagtggctga agatcaagga gcgggcagtg aactctcctg aatcttcgcc tgcttcttca
      481 ttctccttcg tttagctaat agaataactg ctgagttgtg aacagtaagg tgtatgtgag
      541 gtgctcgaaa acaaggtttc aggtgacgcc cccagaataa aatttggacg gggggttcag
      601 tggtggcatt gtgctatgac accaatataa ccctcacaaa ccccttgggc aataaatact
      661 agtgtaggaa tgaaacattc tgaatatctt taacaataga aatccatggg gtggggacaa
      721 gccgtaaaga ctggatgtcc atctcacacg aatttatggc tatgggcaac acataatcct
      781 agtgcaatat gatactgggg ttattaagat gtgtcccagg cagggaccaa gacaggtgaa
      841 ccatgttgtt acactctatt tgtaacaagg ggaaagagag tggacgccga cagcagcgga
      901 ctccactggt tgtctctaac acccccgaaa attaaacggg gctccacgcc aatggggccc
      961 ataaacaaag acaagtggcc actctttttt ttgaaattgt ggagtggggg cacgcgtcag
     1021 cccccacacg ccgccctgcg gttttggact gtaaaataag ggtgtaataa cttggctgat
     1081 tgtaaccccg ctaaccactg cggtcaaacc acttgcccac aaaaccacta atggcacccc
     1141 ggggaatacc tgcataagta ggtgggcggg ccaagatagg ggcgcgattg ctgcgatctg
     1201 gaggacaaat tacacacact tgcgcctgag cgccaagcac agggttgttg gtcctcatat
     1261 tcacgaggtc gctgagagca cggtgggcta atgttgccat gggtagcata tactacccaa
     1321 atatctggat agcatatgct atcctaatct atatctgggt agcataggct atcctaatct
     1381 atatctgggt agcatatgct atcctaatct atatctgggt agtatatgct atcctaattt
     1441 atatctgggt agcataggct atcctaatct atatctgggt agcatatgct atcctaatct
     1501 atatctgggt agtatatgct atcctaatct gtatccgggt agcatatgct atcctaatag
     1561 agattagggt agtatatgct atcctaattt atatctgggt agcatatact acccaaatat
     1621 ctggatagca tatgctatcc taatctatat ctgggtagca tatgctatcc taatctatat
     1681 ctgggtagca taggctatcc taatctatat ctgggtagca tatgctatcc taatctatat
     1741 ctgggtagta tatgctatcc taatttatat ctgggtagca taggctatcc taatctatat
     1801 ctgggtagca tatgctatcc taatctatat ctgggtagta tatgctatcc taatctgtat
     1861 ccgggtagca tatgctatcc tcatgcatat acagtcagca tatgataccc agtagtagag
     1921 tgggagtgct atcctttgca tatgccgcca cctcccaagg gggcgtgaat tttcgctgct
     1981 tgtccttttc ctgctgggga tctcccgatc ccctatggtg cactctcagt acaatctgct
     2041 ctgatgccgc atagttaagc cagtatctgc tccctgcttg tgtgttggag gtcgctgagt
     2101 agtgcgcgag caaaatttaa gctacaacaa ggcaaggctt gaccgacaat tgcatgaaga
     2161 atctgcttag ggttaggcgt tttgcgctgc ttcgatcaca actttggtat agaaaagttg
     2221 gccctgcagg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
     2281 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
     2341 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
     2401 atcatatgcc aagtccgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
     2461 atgcccagta catgacctta cgggactttc ctacttggca gtacatctac gtattagtca
     2521 tcgctattac catggtgatg cggttttggc agtacaccaa tgggcgtgga tagcggtttg
     2581 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
     2641 aaaatcaacg ggactttcca aaatgtcgta ataaccccgc cccgttgacg caaatgggcg
     2701 gtaggcgtgt acggtgggag gtctatataa gcagagctcg tttagtgaac cgtcagatcg
     2761 cctggagacg ccatccacgc tgttttgacc tccatagaag acaacaactt tgtacaaaaa
     2821 agttggcacc atggatgcca tgaagcgggg cctctgttgc gtgctcctcc tgtgcggagc
     2881 tgtgttcgtg agcccttcgg catcgttcag agtgcagccc accgagtcca tcgtgcgctt
     2941 cccgaacatt accaacctct gcccgtttgg ggaggtgttc aatgccacta gattcgcctc
     3001 cgtgtacgcg tggaatcgca agcggatttc caactgcgtc gccgactaca gcgtgctgta
     3061 caactccgcc tccttttcga ccttcaagtg ctacggagtg tcgcccacta agctgaacga
     3121 cctctgcttc accaacgtgt acgcagacag cttcgtgatt aggggggatg aggtccggca
     3181 gatcgctcca gggcagacgg ggaagatcgc cgattacaac tacaagctgc ccgacgactt
     3241 cactggttgc gtcatcgcct ggaactccaa caacctggac agcaaagtcg gcggcaacta
     3301 caactacctg taccggctgt tccgcaagtc caacttgaag ccgttcgagc gggacatcag
     3361 caccgagatc taccaggccg gatcaacccc ctgtaacggc gtcgaaggat tcaactgtta
     3421 cttcccgctg caatcctatg ggtttcagcc gaccaatggc gtgggatatc agccctaccg
     3481 ggtggtggtg ctgtccttcg aacttctgca cgcgccagcc acagtgtgcg gtcctaaaaa
     3541 gggagcgggt tcctccctgg aagtgctgtt ccaaggacct ggctcaggat catcccacca
     3601 ccaccatcat caccaccacg gaggcagcgg atcgtctatg gacgaaaaga ctaccggttg
     3661 gagaggcggc catgtcgtgg aaggcttggc cggagaactg gagcagctta gagccaggct
     3721 ggagcaccac ccgcaaggac agcgcgaacc ttaatcaact ttcttgtaca aagttgtgaa
     3781 attctgcaga tatccatcac actggcggcc gctcgagcat gcatctagag ggccctattc
     3841 tatagtgtca cctaaatgct agagctcgct gatcagcctc gactgtgcct tctagttgcc
     3901 agccatctgt tgtttgcccc tcccccgtgc cttccttgac cctggaaggt gccactccca
     3961 ctgtcctttc ctaataaaat gaggaaattg catcgcattg tctgagtagg tgtcattcta
     4021 ttctgggggg tggggtgggg caggacagca agggggagga ttgggaagac aatagcaggc
     4081 atgctgggga tgcggtgggc tctatggctt ctgaggcgga aagaaccagc tggggctcta
     4141 gggggtatcc ccacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc
     4201 gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt
     4261 cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggc atccctttag
     4321 ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag ggtgatggtt
     4381 cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt
     4441 tctttaatag tggactcttg ttccaaactg gaacaacact caaccctatc tcggtctatt
     4501 cttttgattt ataagggatt ttggggattt cggcctattg gttaaaaaat gagctgattt
     4561 aacaaaaatt taacgcgaat taattctgtg gaatgtgtgt cagttagggt gtggaaagtc
     4621 cccaggctcc ccaggcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca
     4681 ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt
     4741 agtcagcaac catagtcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt
     4801 ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg
     4861 cctctgcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt
     4921 gcaaaaagct cccgggagct tgtatatcca ttttcggatc tgatcaagag acaggatgag
     4981 gatcgtttcg catgattgaa caagatggat tgcacgcagg ttctccggcc gcttgggtgg
     5041 agaggctatt cggctatgac tgggcacaac agacaatcgg ctgctctgat gccgccgtgt
     5101 tccggctgtc agcgcagggg cgcccggttc tttttgtcaa gaccgacctg tccggtgccc
     5161 tgaatgaact gcaggacgag gcagcgcggc tatcgtggct ggccacgacg ggcgttcctt
     5221 gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga ctggctgcta ttgggcgaag
     5281 tgccggggca ggatctcctg tcatctcacc ttgctcctgc cgagaaagta tccatcatgg
     5341 ctgatgcaat gcggcggctg catacgcttg atccggctac ctgcccattc gaccaccaag
     5401 cgaaacatcg catcgagcga gcacgtactc ggatggaagc cggtcttgtc gatcaggatg
     5461 atctggacga agagcatcag gggctcgcgc cagccgaact gttcgccagg ctcaaggcgc
     5521 gcatgcccga cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg ccgaatatca
     5581 tggtggaaaa tggccgcttt tctggattca tcgactgtgg ccggctgggt gtggcggacc
     5641 gctatcagga catagcgttg gctacccgtg atattgctga agagcttggc ggcgaatggg
     5701 ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc atcgccttct
     5761 atcgccttct tgacgagttc ttctgagcgg gactctgggg ttcgaaatga ccgaccaagc
     5821 gacgcccaac ctgccatcac gagatttcga ttccaccgcc gccttctatg aaaggttggg
     5881 cttcggaatc gttttccggg acgccggctg gatgatcctc cagcgcgggg atctcatgct
     5941 ggagttcttc gcccacccca acttgtttat tgcagcttat aatggttaca aataaagcaa
     6001 tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc
     6061 caaactcatc aatgtatctt atcatgtctg tataccgtcg acctctagct agagcttggc
     6121 gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa
     6181 catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac
     6241 attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca
     6301 ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc
     6361 ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc
     6421 aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc
     6481 aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag
     6541 gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc
     6601 gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt
     6661 tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct
     6721 ttctcaatgc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg
     6781 ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct
     6841 tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat
     6901 tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg
     6961 ctacactaga aggacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa
     7021 aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt
     7081 ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc
     7141 tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt
     7201 atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta
     7261 aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat
     7321 ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac
     7381 tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg
     7441 ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag
     7501 tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt
     7561 aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt
     7621 gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt
     7681 tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt
     7741 cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct
     7801 tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt
     7861 ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac
     7921 cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa
     7981 actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa
     8041 ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca
     8101 aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct
     8161 ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga
     8221 atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc
     8281 tgacgtc
//
