LOCUS       Exported                6908 bp ds-DNA     circular SYN 23-AUG-2021
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6908)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Monday, Aug 30, 2021 from SnapGene Viewer 4.3.10
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6908
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     misc_feature    175..317
                     /label=bom
                     /note="basis of mobility region from pBR322"
     CDS             complement(419..610)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(1419..2501)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     promoter        complement(2502..2579)
                     /gene="lacI"
                     /label=lacI promoter
     promoter        2888..2906
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    2907..2931
                     /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)."
     CDS             2982..2999
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             3000..3716
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             3000..3716
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             3720..3740
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             3744..4640
                     /codon_start=1
                     /label=9S [rpoA]
                     /translation="MGNSVVSWNAMISGYAQNGRIDEARELFDKMPERDVVSWNAMISG
                     YAQNGRIDEARELFDKMPERDVVSWTAMISGYAQNGRIDEARELFDKMPERNVVSWNAM
                     ISGYAQNGRIDEARELFDKMPERSVVSWTAMISGYAQNGRIDEARELFDKMPERNVVSW
                     NAMISGYAQNGRIDEARELFDKMPERSVVSWTAMISGYAQNGRIDEARELFDKMPERDV
                     VSWNAMISGYAQNGRIDEARELFDKMPERDVVSWTAMISGYAQNGRIDEARELFDKMPE
                     RDVVSNNAMSAGASQNG"
     CDS             3756..3848
                     /codon_start=1
                     /label=S1
                     /translation="VVSWNAMISGYAQNGRIDEARELFDKMPERD"
     CDS             3849..3941
                     /codon_start=1
                     /label=S2
                     /translation="VVSWNAMISGYAQNGRIDEARELFDKMPERD"
     CDS             3942..4034
                     /codon_start=1
                     /label=S3
                     /translation="VVSWTAMISGYAQNGRIDEARELFDKMPERN"
     CDS             4035..4127
                     /codon_start=1
                     /label=S4
                     /translation="VVSWNAMISGYAQNGRIDEARELFDKMPERS"
     CDS             4128..4220
                     /codon_start=1
                     /label=S5
                     /translation="VVSWTAMISGYAQNGRIDEARELFDKMPERN"
     CDS             4221..4313
                     /codon_start=1
                     /label=S6
                     /translation="VVSWNAMISGYAQNGRIDEARELFDKMPERS"
     CDS             4314..4406
                     /codon_start=1
                     /label=S7
                     /translation="VVSWTAMISGYAQNGRIDEARELFDKMPERD"
     CDS             4407..4499
                     /codon_start=1
                     /label=S8
                     /translation="VVSWNAMISGYAQNGRIDEARELFDKMPERD"
     CDS             4500..4592
                     /codon_start=1
                     /label=S9
                     /translation="VVSWTAMISGYAQNGRIDEARELFDKMPERD"
     CDS             4656..4673
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      4740..4787
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      4824..5279
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     CDS             complement(5372..6187)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     rep_origin      6309..6897
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
       61 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
      121 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcctgat
      181 gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcatatatg gtgcactctc
      241 agtacaatct gctctgatgc cgcatagtta agccagtata cactccgcta tcgctacgtg
      301 actgggtcat ggctgcgccc cgacacccgc caacacccgc tgacgcgccc tgacgggctt
      361 gtctgctccc ggcatccgct tacagacaag ctgtgaccgt ctccgggagc tgcatgtgtc
      421 agaggttttc accgtcatca ccgaaacgcg cgaggcagct gcggtaaagc tcatcagcgt
      481 ggtcgtgaag cgattcacag atgtctgcct gttcatccgc gtccagctcg ttgagtttct
      541 ccagaagcgt taatgtctgg cttctgataa agcgggccat gttaagggcg gttttttcct
      601 gtttggtcac tgatgcctcc gtgtaagggg gatttctgtt catgggggta atgataccga
      661 tgaaacgaga gaggatgctc acgatacggg ttactgatga tgaacatgcc cggttactgg
      721 aacgttgtga gggtaaacaa ctggcggtat ggatgcggcg ggaccagaga aaaatcactc
      781 agggtcaatg ccagcgcttc gttaatacag atgtaggtgt tccacagggt agccagcagc
      841 atcctgcgat gcagatccgg aacataatgg tgcagggcgc tgacttccgc gtttccagac
      901 tttacgaaac acggaaaccg aagaccattc atgttgttgc tcaggtcgca gacgttttgc
      961 agcagcagtc gcttcacgtt cgctcgcgta tcggtgattc attctgctaa ccagtaaggc
     1021 aaccccgcca gcctagccgg gtcctcaacg acaggagcac gatcatgcgc acccgtgggg
     1081 ccgccatgcc ggcgataatg gcctgcttct cgccgaaacg tttggtggcg ggaccagtga
     1141 cgaaggcttg agcgagggcg tgcaagattc cgaataccgc aagcgacagg ccgatcatcg
     1201 tcgcgctcca gcgaaagcgg tcctcgccga aaatgaccca gagcgctgcc ggcacctgtc
     1261 ctacgagttg catgataaag aagacagtca taagtgcggc gacgatagtc atgccccgcg
     1321 cccaccggaa ggagctgact gggttgaagg ctctcaaggg catcggtcga gatcccggtg
     1381 cctaatgagt gagctaactt acattaattg cgttgcgctc actgcccgct ttccagtcgg
     1441 gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc
     1501 gtattgggcg ccagggtggt ttttcttttc accagtgaga cgggcaacag ctgattgccc
     1561 ttcaccgcct ggccctgaga gagttgcagc aagcggtcca cgctggtttg ccccagcagg
     1621 cgaaaatcct gtttgatggt ggttaacggc gggatataac atgagctgtc ttcggtatcg
     1681 tcgtatccca ctaccgagat atccgcacca acgcgcagcc cggactcggt aatggcgcgc
     1741 attgcgccca gcgccatctg atcgttggca accagcatcg cagtgggaac gatgccctca
     1801 ttcagcattt gcatggtttg ttgaaaaccg gacatggcac tccagtcgcc ttcccgttcc
     1861 gctatcggct gaatttgatt gcgagtgaga tatttatgcc agccagccag acgcagacgc
     1921 gccgagacag aacttaatgg gcccgctaac agcgcgattt gctggtgacc caatgcgacc
     1981 agatgctcca cgcccagtcg cgtaccgtct tcatgggaga aaataatact gttgatgggt
     2041 gtctggtcag agacatcaag aaataacgcc ggaacattag tgcaggcagc ttccacagca
     2101 atggcatcct ggtcatccag cggatagtta atgatcagcc cactgacgcg ttgcgcgaga
     2161 agattgtgca ccgccgcttt acaggcttcg acgccgcttc gttctaccat cgacaccacc
     2221 acgctggcac ccagttgatc ggcgcgagat ttaatcgccg cgacaatttg cgacggcgcg
     2281 tgcagggcca gactggaggt ggcaacgcca atcagcaacg actgtttgcc cgccagttgt
     2341 tgtgccacgc ggttgggaat gtaattcagc tccgccatcg ccgcttccac tttttcccgc
     2401 gttttcgcag aaacgtggct ggcctggttc accacgcggg aaacggtctg ataagagaca
     2461 ccggcatact ctgcgacatc gtataacgtt actggtttca cattcaccac cctgaattga
     2521 ctctcttccg ggcgctatca tgccataccg cgaaaggttt tgcgccattc gatggtgtcc
     2581 gggatctcga cgctctccct tatgcgactc ctgcattagg aagcagccca gtagtaggtt
     2641 gaggccgttg agcaccgccg ccgcaaggaa tggtgcatgc aaggagatgg cgcccaacag
     2701 tcccccggcc acggggcctg ccaccatacc cacgccgaaa caagcgctca tgagcccgaa
     2761 gtggcgagcc cgatcttccc catcggtgat gtcggcgata taggcgccag caaccgcacc
     2821 tgtggcgccg gtgatgccgg ccacgatgcg tccggcgtag aggatcgaga tctcgatccc
     2881 gcgaaattaa tacgactcac tataggggaa ttgtgagcgg ataacaattc ccctctagaa
     2941 ataattttga tttaacttta agaaggagat ataccatgaa acatcaccat caccatcaca
     3001 tggtgagcaa gggcgaggag ctgttcaccg gggtggtgcc catcctggtc gagctggacg
     3061 gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg cgagggcgat gccacctacg
     3121 gcaagctgac cctgaagttc atctgcacca ccggcaagct gcccgtgccc tggcccaccc
     3181 tcgtgaccac cctgacctac ggcgtgcagt gcttcagccg ctaccccgac cacatgaagc
     3241 agcacgactt cttcaagtcc gccatgcccg aaggctacgt ccaggagcgc accatcttct
     3301 tcaaggacga cggcaactac aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg
     3361 tgaaccgcat cgagctgaag ggcatcgact tcaaggagga cggcaacatc ctggggcaca
     3421 agctggagta caactacaac agccacaacg tctatatcat ggccgacaag cagaagaacg
     3481 gcatcaaggt gaacttcaag atccgccaca acatcgagga cggcagcgtg cagctcgccg
     3541 accactacca gcagaacacc cccatcggcg acggccccgt gctgctgccc gacaaccact
     3601 acctgagcac ccagtccgcc ctgagcaaag accccaacga gaagcgcgat cacatggtcc
     3661 tgctggagtt cgtgaccgcc gccgggatca ctctcggcat ggacgagctg tacaaggcgg
     3721 agaatcttta ttttcagggc gccatgggaa attctgtggt gtcgtggaac gctatgatca
     3781 gcgggtacgc gcaaaacggc cggatcgatg aagcgcgaga actatttgat aagatgccgg
     3841 agagagacgt tgtctcctgg aatgcgatga tttcaggata tgcgcagaac gggaggattg
     3901 atgaggctcg ggagttattt gacaaaatgc cagagcggga cgtggtttct tggactgcca
     3961 tgatctccgg gtacgcacaa aacggacgca ttgacgaagc gcgggagctc tttgataaaa
     4021 tgcccgagcg caacgtcgtg tcttggaatg ccatgatttc ggggtatgcc caaaatggtc
     4081 gcatcgacga ggcgcgagaa ttattcgaca agatgccaga acggtcggtg gtgtcctgga
     4141 ccgctatgat tagtggctac gcccaaaacg ggcgaataga tgaagcccgt gaactatttg
     4201 acaagatgcc tgaacgcaat gtcgtgtcgt ggaatgcaat gatctcaggg tatgcgcaga
     4261 atggacgaat agacgaagcg cgagagctgt tcgataaaat gccggaacga tcggtagtat
     4321 catggacggc aatgatatcg gggtacgctc aaaatggtcg gattgatgag gcacgcgaac
     4381 tgttcgacaa aatgcctgag agggatgtcg tctcatggaa cgcgatgatc tccggatatg
     4441 ctcaaaatgg ccggattgac gaggcgagag aactcttcga taagatgccc gagcgggacg
     4501 tcgtttcgtg gacagccatg atcagtggtt acgcacaaaa cgggcgcatc gatgaagcta
     4561 gggaactgtt cgacaagatg cccgaaaggg atgtagtctc gaacaacgcc atgtcggctg
     4621 gagcgtcgca gaatggctaa gcggccgcac tcgagcacca ccaccaccac cactgagatc
     4681 cggctgctaa caaagcccga aaggaagctg agttggctgc tgccaccgct gagcaataac
     4741 tagcataacc ccttggggcc tctaaacggg tcttgagggg ttttttgctg aaaggaggaa
     4801 ctatatccgg attggcgaat gggacgcgcc ctgtagcggc gcattaagcg cggcgggtgt
     4861 ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc
     4921 tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg
     4981 gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta
     5041 gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt
     5101 ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat
     5161 ctcggtctat tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa
     5221 tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa atattaacgt ttacaatttc
     5281 aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca
     5341 ttcaaatatg tatccgctca tgaattaatt cttagaaaaa ctcatcgagc atcaaatgaa
     5401 actgcaattt attcatatca ggattatcaa taccatattt ttgaaaaagc cgtttctgta
     5461 atgaaggaga aaactcaccg aggcagttcc ataggatggc aagatcctgg tatcggtctg
     5521 cgattccgac tcgtccaaca tcaatacaac ctattaattt cccctcgtca aaaataaggt
     5581 tatcaagtga gaaatcacca tgagtgacga ctgaatccgg tgagaatggc aaaagtttat
     5641 gcatttcttt ccagacttgt tcaacaggcc agccattacg ctcgtcatca aaatcactcg
     5701 catcaaccaa accgttattc attcgtgatt gcgcctgagc gagacgaaat acgcgatcgc
     5761 tgttaaaagg acaattacaa acaggaatcg aatgcaaccg gcgcaggaac actgccagcg
     5821 catcaacaat attttcacct gaatcaggat attcttctaa tacctggaat gctgttttcc
     5881 cggggatcgc agtggtgagt aaccatgcat catcaggagt acggataaaa tgcttgatgg
     5941 tcggaagagg cataaattcc gtcagccagt ttagtctgac catctcatct gtaacatcat
     6001 tggcaacgct acctttgcca tgtttcagaa acaactctgg cgcatcgggc ttcccataca
     6061 atcgatagat tgtcgcacct gattgcccga cattatcgcg agcccattta tacccatata
     6121 aatcagcatc catgttggaa tttaatcgcg gcctagagca agacgtttcc cgttgaatat
     6181 ggctcataac accccttgta ttactgttta tgtaagcaga cagttttatt gttcatgacc
     6241 aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa
     6301 ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca
     6361 ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta
     6421 actggcttca gcagagcgca gataccaaat actgtccttc tagtgtagcc gtagttaggc
     6481 caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca
     6541 gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta
     6601 ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag
     6661 cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt
     6721 cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc
     6781 acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac
     6841 ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac
     6901 gccagcaa
//
