LOCUS       Exported                6730 bp ds-DNA     circular SYN 12-OCT-2022
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6730)
  AUTHORS   MPIB
  TITLE     Direct Submission
  JOURNAL   Exported Friday, Jan 19, 2024 from SnapGene 5.0.8
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6730
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     promoter        21..39
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    40..64
                     /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)."
     RBS             79..101
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             120..137
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             147..164
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             168..389
                     /codon_start=1
                     /product="human ubiquitin"
                     /label=ubiquitin
                     /translation="MQIFVKTLTGKTITLEVEPSDTIENVKAKIQDKEGIPPDQQRLIF
                     AGKQLEDGRTLSDYNIQKESTLHLVLRLR"
     primer_bind     386..447
                     /label=Primer 1
     CDS             429..1147
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1124..1224)
                     /label=Primer 2
     terminator      1289..1336
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        1625..1729
                     /gene="bla"
                     /label=AmpR promoter
     CDS             1730..2590
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      2761..3349
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     misc_feature    3535..3677
                     /label=bom
                     /note="basis of mobility region from pBR322"
     CDS             complement(3779..3970)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(5282..6364)
                     /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(6365..6442)
                     /gene="lacI"
                     /label=lacI promoter
ORIGIN
        1 agatctcgat cccgcgaaat taatacgact cactataggg gaattgtgag cggataacaa
       61 ttcccctcta gaaataattt tgtttaactt taagaaggag atataccatg ggcagcagcc
      121 atcatcatca tcatcacagc agcggcctgg tgccgcgcgg cagccatatg cagatctttg
      181 tgaagaccct cactggcaaa accatcaccc ttgaggtcga gcccagtgac accattgaga
      241 atgtcaaagc caaaattcaa gacaaggagg gtatcccacc tgaccagcag cgtctgatat
      301 ttgccggcaa acagctggag gatggccgca ctctctcaga ctacaacatc cagaaagagt
      361 ccaccctgca cctggtgttg cgcctccgcg gtggtagctt gatgcccttc tctccgtacg
      421 agcccctgat ggtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg
      481 agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg
      541 ccacctacgg caagctgacc ctgaagttca tctgcaccac cggcaagctg cccgtgccct
      601 ggcccaccct cgtgaccacc ctgacctacg gcgtgcagtg cttcagccgc taccccgacc
      661 acatgaagca gcacgacttc ttcaagtccg ccatgcccga aggctacgtc caggagcgca
      721 ccatcttctt caaggacgac ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg
      781 acaccctggt gaaccgcatc gagctgaagg gcatcgactt caaggaggac ggcaacatcc
      841 tggggcacaa gctggagtac aactacaaca gccacaacgt ctatatcatg gccgacaagc
      901 agaagaacgg catcaaggtg aacttcaaga tccgccacaa catcgaggac ggcagcgtgc
      961 agctcgccga ccactaccag cagaacaccc ccatcggcga cggccccgtg ctgctgcccg
     1021 acaaccacta cctgagcacc cagtccgccc tgagcaaaga ccccaacgag aagcgcgatc
     1081 acatggtcct gctggagttc gtgaccgccg ccgggatcac tctcggcatg gacgagctgt
     1141 acaagtacca ctacctgccc ttcagcctgc cccaccggag gcctcacttc ttctttccca
     1201 agtcccgcat cgtccgctaa gcttagatcc ggctgctaac aaagcccgaa aggaagctga
     1261 gttggctgct gccaccgctg agcaataact agcataaccc cttggggcct ctaaacgggt
     1321 cttgaggggt tttttgctga aaggaggaac tatatccgga tatcccgcaa gaggcccggc
     1381 agtaccggca taaccaagcc tatgcctaca gcatccaggg tgacggtgcc gaggatgacg
     1441 atgagcgcat tgttagattt catacacggt gcctgactgc gttagcaatt taactgtgat
     1501 aaactaccgc attaaagcta attcttgaag acgaaagggc ctcgtgatac gcctattttt
     1561 ataggttaat gtcatgataa taatggtttc ttagacgtca ggtggcactt ttcggggaaa
     1621 tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat
     1681 gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca
     1741 acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca
     1801 cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta
     1861 catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt
     1921 tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc gtgttgacgc
     1981 cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc
     2041 accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc
     2101 cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa
     2161 ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga
     2221 accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgcagcaat
     2281 ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca
     2341 attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc
     2401 ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat
     2461 tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag
     2521 tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa
     2581 gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca
     2641 tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc
     2701 ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc
     2761 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
     2821 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
     2881 cagcagagcg cagataccaa atactgtcct tctagtgtag ccgtagttag gccaccactt
     2941 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
     3001 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
     3061 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
     3121 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
     3181 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
     3241 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
     3301 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
     3361 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
     3421 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
     3481 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcctgat
     3541 gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcatatatg gtgcactctc
     3601 agtacaatct gctctgatgc cgcatagtta agccagtata cactccgcta tcgctacgtg
     3661 actgggtcat ggctgcgccc cgacacccgc caacacccgc tgacgcgccc tgacgggctt
     3721 gtctgctccc ggcatccgct tacagacaag ctgtgaccgt ctccgggagc tgcatgtgtc
     3781 agaggttttc accgtcatca ccgaaacgcg cgaggcagct gcggtaaagc tcatcagcgt
     3841 ggtcgtgaag cgattcacag atgtctgcct gttcatccgc gtccagctcg ttgagtttct
     3901 ccagaagcgt taatgtctgg cttctgataa agcgggccat gttaagggcg gttttttcct
     3961 gtttggtcac tgatgcctcc gtgtaagggg gatttctgtt catgggggta atgataccga
     4021 tgaaacgaga gaggatgctc acgatacggg ttactgatga tgaacatgcc cggttactgg
     4081 aacgttgtga gggtaaacaa ctggcggtat ggatgcggcg ggaccagaga aaaatcactc
     4141 agggtcaatg ccagcgcttc gttaatacag atgtaggtgt tccacagggt agccagcagc
     4201 atcctgcgat gcagatccgg aacataatgg tgcagggcgc tgacttccgc gtttccagac
     4261 tttacgaaac acggaaaccg aagaccattc atgttgttgc tcaggtcgca gacgttttgc
     4321 agcagcagtc gcttcacgtt cgctcgcgta tcggtgattc attctgctaa ccagtaaggc
     4381 aaccccgcca gcctagccgg gtcctcaacg acaggagcac gatcatgcgc acccgtggcc
     4441 aggacccaac gctgcccgag atgcgccgcg tgcggctgct ggagatggcg gacgcgatgg
     4501 atatgttctg ccaagggttg gtttgcgcat tcacagttct ccgcaagaat tgattggctc
     4561 caattcttgg agtggtgaat ccgttagcga ggtgccgccg gcttccattc aggtcgaggt
     4621 ggcccggctc catgcaccgc gacgcaacgc ggggaggcag acaaggtata gggcggcgcc
     4681 tacaatccat gccaacccgt tccatgtgct cgccgaggcg gcataaatcg ccgtgacgat
     4741 cagcggtcca gtgatcgaag ttaggctggt aagagccgcg agcgatcctt gaagctgtcc
     4801 ctgatggtcg tcatctacct gcctggacag catggcctgc aacgcgggca tcccgatgcc
     4861 gccggaagcg agaagaatca taatggggaa ggccatccag cctcgcgtcg cgaacgccag
     4921 caagacgtag cccagcgcgt cggccgccat gccggcgata atggcctgct tctcgccgaa
     4981 acgtttggtg gcgggaccag tgacgaaggc ttgagcgagg gcgtgcaaga ttccgaatac
     5041 cgcaagcgac aggccgatca tcgtcgcgct ccagcgaaag cggtcctcgc cgaaaatgac
     5101 ccagagcgct gccggcacct gtcctacgag ttgcatgata aagaagacag tcataagtgc
     5161 ggcgacgata gtcatgcccc gcgcccaccg gaaggagctg actgggttga aggctctcaa
     5221 gggcatcggt cgagatcccg gtgcctaatg agtgagctaa cttacattaa ttgcgttgcg
     5281 ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca
     5341 acgcgcgggg agaggcggtt tgcgtattgg gcgccagggt ggtttttctt ttcaccagtg
     5401 agacgggcaa cagctgattg cccttcaccg cctggccctg agagagttgc agcaagcggt
     5461 ccacgctggt ttgccccagc aggcgaaaat cctgtttgat ggtggttaac ggcgggatat
     5521 aacatgagct gtcttcggta tcgtcgtatc ccactaccga gatatccgca ccaacgcgca
     5581 gcccggactc ggtaatggcg cgcattgcgc ccagcgccat ctgatcgttg gcaaccagca
     5641 tcgcagtggg aacgatgccc tcattcagca tttgcatggt ttgttgaaaa ccggacatgg
     5701 cactccagtc gccttcccgt tccgctatcg gctgaatttg attgcgagtg agatatttat
     5761 gccagccagc cagacgcaga cgcgccgaga cagaacttaa tgggcccgct aacagcgcga
     5821 tttgctggtg acccaatgcg accagatgct ccacgcccag tcgcgtaccg tcttcatggg
     5881 agaaaataat actgttgatg ggtgtctggt cagagacatc aagaaataac gccggaacat
     5941 tagtgcaggc agcttccaca gcaatggcat cctggtcatc cagcggatag ttaatgatca
     6001 gcccactgac gcgttgcgcg agaagattgt gcaccgccgc tttacaggct tcgacgccgc
     6061 ttcgttctac catcgacacc accacgctgg cacccagttg atcggcgcga gatttaatcg
     6121 ccgcgacaat ttgcgacggc gcgtgcaggg ccagactgga ggtggcaacg ccaatcagca
     6181 acgactgttt gcccgccagt tgttgtgcca cgcggttggg aatgtaattc agctccgcca
     6241 tcgccgcttc cactttttcc cgcgttttcg cagaaacgtg gctggcctgg ttcaccacgc
     6301 gggaaacggt ctgataagag acaccggcat actctgcgac atcgtataac gttactggtt
     6361 tcacattcac caccctgaat tgactctctt ccgggcgcta tcatgccata ccgcgaaagg
     6421 ttttgcgcca ttcgatggtg tccgggatct cgacgctctc ccttatgcga ctcctgcatt
     6481 aggaagcagc ccagtagtag gttgaggccg ttgagcaccg ccgccgcaag gaatggtgca
     6541 tgcaaggaga tggcgcccaa cagtcccccg gccacggggc ctgccaccat acccacgccg
     6601 aaacaagcgc tcatgagccc gaagtggcga gcccgatctt ccccatcggt gatgtcggcg
     6661 atataggcgc cagcaaccgc acctgtggcg ccggtgatgc cggccacgat gcgtccggcg
     6721 tagaggatcg
//
