LOCUS       40924_17926        5821 bp DNA     circular SYN 13-JAN-2022
DEFINITION  synthetic circular DNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5821)
  AUTHORS   caoheibi
  TITLE     Direct Submission
REFERENCE   2  (bases 1 to 5821)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..5821
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(26..73)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(150..173)
                     /codon_start=1
                     /label=8xHis
                     /note="8xHis affinity tag"
                     /translation="HHHHHHHH"
     CDS             complement(528..545)
                     /codon_start=1
                     /label=thrombin site
                     /note="thrombin recognition and cleavage site"
                     /translation="LVPRGS"
     RBS             complement(754..776)
                     /label=RBS
                     /note="efficient ribosome binding site from bacteriophage
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    complement(791..815)
                     /label=lac operator
                     /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(816..834)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        1147..1224
                     /label=lacI promoter
     CDS             1225..2304
                     /codon_start=1
                     /label=lacI
                     /note="lac repressor"
                     /translation="VKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     protein_bind    2320..2341
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     CDS             3116..3304
                     /codon_start=1
                     /label=rop
                     /note="Rop protein, which maintains plasmids at low copy
                     number"
                     /translation="VTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     misc_feature    3409..3551
                     /label=bom
                     /note="basis of mobility region from pBR322"
     rep_origin      complement(3737..4325)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             4447..5259
                     /codon_start=1
                     /label=KanR
                     /note="aminoglycoside phosphotransferase"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     rep_origin      complement(5355..5810)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
ORIGIN
        1 atccggatat agttcctcct ttcagcaaaa aacccctcaa gacccgttta gaggccccaa
       61 ggggttatgc tagttattgc tcagcggtgg cagcagccaa ctcagcttcc tttcgggctt
      121 tgtttagcag cctaggtatt aatcaattag tggtggtggt ggtggtggtg gtgctcgacg
      181 ccgaccgtgc agggcgtgcc gttgagcgag aacgccgtcg gcgcggcgtt ggtgcccgtg
      241 tgcgagccgt tgaagccgaa ctgcgcggtg ccgcccgccg ggatcgagcc gttccacggg
      301 gcgttgcgga ccgtcacggc cgagccggac tgcgtgaccg tcgagctcca cgcctgggtg
      361 acctgctggc cggacgggaa gctgaacgtg agcgtccagc cgtctaccgg agcggaggac
      421 gtgttcttca cggtgacgtt cgcggtgaag ccggtgttcc actggttgac gccccacagc
      481 acctggcacc cggccggacc actagtcggg gtcggcgtcg gaggaccgga accgcgtggc
      541 accagaccct cgagtgcggc cgcaagcttg acgtcctgca ggtgtacaga attcggatcc
      601 gatatcacca tggtgctagc tgcctgagct ggcagaacaa cagttgcacc tacaaccagg
      661 gtagcagcag cagcagccag agtagtacgc agagcagtac gagcaccacg agccggatga
      721 cctggagctg gagtagtacg aggcatatgt atatctcctt cttaaagtta aacaaaatta
      781 tttctagagg ggaattgtta tccgctcaca attcccctat agtgagtcgt attaatttcg
      841 cgggatcgag atcgatctcg atcctctacg ccggacgcat cgtggccggc atcaccggcg
      901 ccacaggtgc ggttgctggc gcctatatcg ccgacatcac cgatggggaa gatcgggctc
      961 gccacttcgg gctcatgagc gcttgtttcg gcgtgggtat ggtggcaggc cccgtggccg
     1021 ggggactgtt gggcgccatc tccttgcatg caccattcct tgcggcggcg gtgctcaacg
     1081 gcctcaacct actactgggc tgcttcctaa tgcaggagtc gcataaggga gagcgtcgag
     1141 atcccggaca ccatcgaatg gcgcaaaacc tttcgcggta tggcatgata gcgcccggaa
     1201 gagagtcaat tcagggtggt gaatgtgaaa ccagtaacgt tatacgatgt cgcagagtat
     1261 gccggtgtct cttatcagac cgtttcccgc gtggtgaacc aggccagcca cgtttctgcg
     1321 aaaacgcggg aaaaagtgga agcggcgatg gcggagctga attacattcc caaccgcgtg
     1381 gcacaacaac tggcgggcaa acagtcgttg ctgattggcg ttgccacctc cagtctggcc
     1441 ctgcacgcgc cgtcgcaaat tgtcgcggcg attaaatctc gcgccgatca actgggtgcc
     1501 agcgtggtgg tgtcgatggt agaacgaagc ggcgtcgaag cctgtaaagc ggcggtgcac
     1561 aatcttctcg cgcaacgcgt cagtgggctg atcattaact atccgctgga tgaccaggat
     1621 gccattgctg tggaagctgc ctgcactaat gttccggcgt tatttcttga tgtctctgac
     1681 cagacaccca tcaacagtat tattttctcc catgaagacg gtacgcgact gggcgtggag
     1741 catctggtcg cattgggtca ccagcaaatc gcgctgttag cgggcccatt aagttctgtc
     1801 tcggcgcgtc tgcgtctggc tggctggcat aaatatctca ctcgcaatca aattcagccg
     1861 atagcggaac gggaaggcga ctggagtgcc atgtccggtt ttcaacaaac catgcaaatg
     1921 ctgaatgagg gcatcgttcc cactgcgatg ctggttgcca acgatcagat ggcgctgggc
     1981 gcaatgcgcg ccattaccga gtccgggctg cgcgttggtg cggacatctc ggtagtggga
     2041 tacgacgata ccgaagacag ctcatgttat atcccgccgt taaccaccat caaacaggat
     2101 tttcgcctgc tggggcaaac cagcgtggac cgcttgctgc aactctctca gggccaggcg
     2161 gtgaagggca atcagctgtt gcccgtctca ctggtgaaaa gaaaaaccac cctggcgccc
     2221 aatacgcaaa ccgcctctcc ccgcgcgttg gccgattcat taatgcagct ggcacgacag
     2281 gtttcccgac tggaaagcgg gcagtgagcg caacgcaatt aatgtaagtt agctcactca
     2341 ttaggcaccg ggatctcgac cgatgccctt gagagccttc aacccagtca gctccttccg
     2401 gtgggcgcgg ggcatgacta tcgtcgccgc acttatgact gtcttcttta tcatgcaact
     2461 cgtaggacag gtgccggcag cgctctgggt cattttcggc gaggaccgct ttcgctggag
     2521 cgcgacgatg atcggcctgt cgcttgcggt attcggaatc ttgcacgccc tcgctcaagc
     2581 cttcgtcact ggtcccgcca ccaaacgttt cggcgagaag caggccatta tcgccggcat
     2641 ggcggcccca cgggtgcgca tgatcgtgct cctgtcgttg aggacccggc taggctggcg
     2701 gggttgcctt actggttagc agaatgaatc accgatacgc gagcgaacgt gaagcgactg
     2761 ctgctgcaaa acgtctgcga cctgagcaac aacatgaatg gtcttcggtt tccgtgtttc
     2821 gtaaagtctg gaaacgcgga agtcagcgcc ctgcaccatt atgttccgga tctgcatcgc
     2881 aggatgctgc tggctaccct gtggaacacc tacatctgta ttaacgaagc gctggcattg
     2941 accctgagtg atttttctct ggtcccgccg catccatacc gccagttgtt taccctcaca
     3001 acgttccagt aaccgggcat gttcatcatc agtaacccgt atcgtgagca tcctctctcg
     3061 tttcatcggt atcattaccc ccatgaacag aaatccccct tacacggagg catcagtgac
     3121 caaacaggaa aaaaccgccc ttaacatggc ccgctttatc agaagccaga cattaacgct
     3181 tctggagaaa ctcaacgagc tggacgcgga tgaacaggca gacatctgtg aatcgcttca
     3241 cgaccacgct gatgagcttt accgcagctg cctcgcgcgt ttcggtgatg acggtgaaaa
     3301 cctctgacac atgcagctcc cggagacggt cacagcttgt ctgtaagcgg atgccgggag
     3361 cagacaagcc cgtcagggcg cgtcagcggg tgttggcggg tgtcggggcg cagccatgac
     3421 ccagtcacgt agcgatagcg gagtgtatac tggcttaact atgcggcatc agagcagatt
     3481 gtactgagag tgcaccatat atgcggtgtg aaataccgca cagatgcgta aggagaaaat
     3541 accgcatcag gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc
     3601 tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg
     3661 ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg
     3721 ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac
     3781 gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg
     3841 gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct
     3901 ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg
     3961 tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct
     4021 gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac
     4081 tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt
     4141 tcttgaagtg gtggcctaac tacggctaca ctagaaggac agtatttggt atctgcgctc
     4201 tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca
     4261 ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat
     4321 ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac
     4381 gttaagggat tttggtcatg aacaataaaa ctgtctgctt acataaacag taatacaagg
     4441 ggtgttatga gccatattca acgggaaacg tcttgctcta ggccgcgatt aaattccaac
     4501 atggatgctg atttatatgg gtataaatgg gctcgcgata atgtcgggca atcaggtgcg
     4561 acaatctatc gattgtatgg gaagcccgat gcgccagagt tgtttctgaa acatggcaaa
     4621 ggtagcgttg ccaatgatgt tacagatgag atggtcagac taaactggct gacggaattt
     4681 atgcctcttc cgaccatcaa gcattttatc cgtactcctg atgatgcatg gttactcacc
     4741 actgcgatcc ccgggaaaac agcattccag gtattagaag aatatcctga ttcaggtgaa
     4801 aatattgttg atgcgctggc agtgttcctg cgccggttgc attcgattcc tgtttgtaat
     4861 tgtcctttta acagcgatcg cgtatttcgt ctcgctcagg cgcaatcacg aatgaataac
     4921 ggtttggttg atgcgagtga ttttgatgac gagcgtaatg gctggcctgt tgaacaagtc
     4981 tggaaagaaa tgcataaact tttgccattc tcaccggatt cagtcgtcac tcatggtgat
     5041 ttctcacttg ataaccttat ttttgacgag gggaaattaa taggttgtat tgatgttgga
     5101 cgagtcggaa tcgcagaccg ataccaggat cttgccatcc tatggaactg cctcggtgag
     5161 ttttctcctt cattacagaa acggcttttt caaaaatatg gtattgataa tcctgatatg
     5221 aataaattgc agtttcattt gatgctcgat gagtttttct aagaattaat tcatgagcgg
     5281 atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg
     5341 aaaagtgcca cctgaaattg taaacgttaa tattttgtta aaattcgcgt taaatttttg
     5401 ttaaatcagc tcatttttta accaataggc cgaaatcggc aaaatccctt ataaatcaaa
     5461 agaatagacc gagatagggt tgagtgttgt tccagtttgg aacaagagtc cactattaaa
     5521 gaacgtggac tccaacgtca aagggcgaaa aaccgtctat cagggcgatg gcccactacg
     5581 tgaaccatca ccctaatcaa gttttttggg gtcgaggtgc cgtaaagcac taaatcggaa
     5641 ccctaaaggg agcccccgat ttagagcttg acggggaaag ccggcgaacg tggcgagaaa
     5701 ggaagggaag aaagcgaaag gagcgggcgc tagggcgctg gcaagtgtag cggtcacgct
     5761 gcgcgtaacc accacacccg ccgcgcttaa tgcgccgcta cagggcgcgt cccattcgcc
     5821 a
//
