LOCUS       40924_17801        5633 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 5633)
  AUTHORS   caoheibi
  TITLE     Direct Submission
REFERENCE   2  (bases 1 to 5633)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..5633
                     /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(140..157)
                     /codon_start=1
                     /label=6xHis
                     /note="6xHis affinity tag"
                     /translation="HHHHHH"
     CDS             complement(201..494)
                     /codon_start=1
                     /label=SUMO
                     /note="cleavable ubiquitin-like protein tag"
                     /translation="MSDSEVNQEAKPEVKPEVKPETHINLKVSDGSSEIFFKIKKTTPL
                     RRLMEAFAKRQGKEMDSLRFLYDGIRIQADQTPEDLDMEDNDIIEAHREQIGG"
     CDS             complement(507..524)
                     /codon_start=1
                     /label=thrombin site
                     /note="thrombin recognition and cleavage site"
                     /translation="LVPRGS"
     CDS             complement(534..551)
                     /codon_start=1
                     /label=6xHis
                     /note="6xHis affinity tag"
                     /translation="HHHHHH"
     RBS             complement(570..592)
                     /label=RBS
                     /note="efficient ribosome binding site from bacteriophage
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    complement(607..631)
                     /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(632..650)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        959..1036
                     /label=lacI promoter
     CDS             1037..2116
                     /codon_start=1
                     /label=lacI
                     /note="lac repressor"
                     /translation="VKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     protein_bind    2132..2153
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     CDS             2928..3116
                     /codon_start=1
                     /label=rop
                     /note="Rop protein, which maintains plasmids at low copy
                     number"
                     /translation="VTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     misc_feature    3221..3363
                     /label=bom
                     /note="basis of mobility region from pBR322"
     rep_origin      complement(3549..4137)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             4259..5071
                     /codon_start=1
                     /label=KanR
                     /note="aminoglycoside phosphotransferase"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     rep_origin      complement(5167..5622)
                     /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 tgttagcagc cggatctcag tggtggtggt ggtggtgctc gagtgcggcc gcaagcttgt
      181 cgacggagct cgaattcgga tccaccaatc tgttctctgt gagcctcaat aatatcgtta
      241 tcctccatgt ccaaatcttc aggggtctga tcagcttgaa ttctaatacc gtcgtacaag
      301 aatcttaagg agtccatttc cttaccctgt cttttagcga acgcttccat cagccttctt
      361 aaaggagtgg tctttttgat cttgaagaag atctctgaag atccatcgga cacctttaaa
      421 ttgatgtgag tctcaggctt gacttctggc ttgacctctg gcttagcttc ttgattgact
      481 tctgagtccg acatgctagc catatggctg ccgcgcggca ccaggccgct gctgtgatga
      541 tgatgatgat ggctgctgcc catggtatat ctccttctta aagttaaaca aaattatttc
      601 tagaggggaa ttgttatccg ctcacaattc ccctatagtg agtcgtatta atttcgcggg
      661 atcgagatct cgatcctcta cgccggacgc atcgtggccg gcatcaccgg cgccacaggt
      721 gcggttgctg gcgcctatat cgccgacatc accgatgggg aagatcgggc tcgccacttc
      781 gggctcatga gcgcttgttt cggcgtgggt atggtggcag gccccgtggc cgggggactg
      841 ttgggcgcca tctccttgca tgcaccattc cttgcggcgg cggtgctcaa cggcctcaac
      901 ctactactgg gctgcttcct aatgcaggag tcgcataagg gagagcgtcg agatcccgga
      961 caccatcgaa tggcgcaaaa cctttcgcgg tatggcatga tagcgcccgg aagagagtca
     1021 attcagggtg gtgaatgtga aaccagtaac gttatacgat gtcgcagagt atgccggtgt
     1081 ctcttatcag accgtttccc gcgtggtgaa ccaggccagc cacgtttctg cgaaaacgcg
     1141 ggaaaaagtg gaagcggcga tggcggagct gaattacatt cccaaccgcg tggcacaaca
     1201 actggcgggc aaacagtcgt tgctgattgg cgttgccacc tccagtctgg ccctgcacgc
     1261 gccgtcgcaa attgtcgcgg cgattaaatc tcgcgccgat caactgggtg ccagcgtggt
     1321 ggtgtcgatg gtagaacgaa gcggcgtcga agcctgtaaa gcggcggtgc acaatcttct
     1381 cgcgcaacgc gtcagtgggc tgatcattaa ctatccgctg gatgaccagg atgccattgc
     1441 tgtggaagct gcctgcacta atgttccggc gttatttctt gatgtctctg accagacacc
     1501 catcaacagt attattttct cccatgaaga cggtacgcga ctgggcgtgg agcatctggt
     1561 cgcattgggt caccagcaaa tcgcgctgtt agcgggccca ttaagttctg tctcggcgcg
     1621 tctgcgtctg gctggctggc ataaatatct cactcgcaat caaattcagc cgatagcgga
     1681 acgggaaggc gactggagtg ccatgtccgg ttttcaacaa accatgcaaa tgctgaatga
     1741 gggcatcgtt cccactgcga tgctggttgc caacgatcag atggcgctgg gcgcaatgcg
     1801 cgccattacc gagtccgggc tgcgcgttgg tgcggatatc tcggtagtgg gatacgacga
     1861 taccgaagac agctcatgtt atatcccgcc gttaaccacc atcaaacagg attttcgcct
     1921 gctggggcaa accagcgtgg accgcttgct gcaactctct cagggccagg cggtgaaggg
     1981 caatcagctg ttgcccgtct cactggtgaa aagaaaaacc accctggcgc ccaatacgca
     2041 aaccgcctct ccccgcgcgt tggccgattc attaatgcag ctggcacgac aggtttcccg
     2101 actggaaagc gggcagtgag cgcaacgcaa ttaatgtaag ttagctcact cattaggcac
     2161 cgggatctcg accgatgccc ttgagagcct tcaacccagt cagctccttc cggtgggcgc
     2221 ggggcatgac tatcgtcgcc gcacttatga ctgtcttctt tatcatgcaa ctcgtaggac
     2281 aggtgccggc agcgctctgg gtcattttcg gcgaggaccg ctttcgctgg agcgcgacga
     2341 tgatcggcct gtcgcttgcg gtattcggaa tcttgcacgc cctcgctcaa gccttcgtca
     2401 ctggtcccgc caccaaacgt ttcggcgaga agcaggccat tatcgccggc atggcggccc
     2461 cacgggtgcg catgatcgtg ctcctgtcgt tgaggacccg gctaggctgg cggggttgcc
     2521 ttactggtta gcagaatgaa tcaccgatac gcgagcgaac gtgaagcgac tgctgctgca
     2581 aaacgtctgc gacctgagca acaacatgaa tggtcttcgg tttccgtgtt tcgtaaagtc
     2641 tggaaacgcg gaagtcagcg ccctgcacca ttatgttccg gatctgcatc gcaggatgct
     2701 gctggctacc ctgtggaaca cctacatctg tattaacgaa gcgctggcat tgaccctgag
     2761 tgatttttct ctggtcccgc cgcatccata ccgccagttg tttaccctca caacgttcca
     2821 gtaaccgggc atgttcatca tcagtaaccc gtatcgtgag catcctctct cgtttcatcg
     2881 gtatcattac ccccatgaac agaaatcccc cttacacgga ggcatcagtg accaaacagg
     2941 aaaaaaccgc ccttaacatg gcccgcttta tcagaagcca gacattaacg cttctggaga
     3001 aactcaacga gctggacgcg gatgaacagg cagacatctg tgaatcgctt cacgaccacg
     3061 ctgatgagct ttaccgcagc tgcctcgcgc gtttcggtga tgacggtgaa aacctctgac
     3121 acatgcagct cccggagacg gtcacagctt gtctgtaagc ggatgccggg agcagacaag
     3181 cccgtcaggg cgcgtcagcg ggtgttggcg ggtgtcgggg cgcagccatg acccagtcac
     3241 gtagcgatag cggagtgtat actggcttaa ctatgcggca tcagagcaga ttgtactgag
     3301 agtgcaccat atatgcggtg tgaaataccg cacagatgcg taaggagaaa ataccgcatc
     3361 aggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga
     3421 gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca
     3481 ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg
     3541 ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt
     3601 cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc
     3661 ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct
     3721 tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc
     3781 gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta
     3841 tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca
     3901 gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag
     3961 tggtggccta actacggcta cactagaagg acagtatttg gtatctgcgc tctgctgaag
     4021 ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt
     4081 agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa
     4141 gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg
     4201 attttggtca tgaacaataa aactgtctgc ttacataaac agtaatacaa ggggtgttat
     4261 gagccatatt caacgggaaa cgtcttgctc taggccgcga ttaaattcca acatggatgc
     4321 tgatttatat gggtataaat gggctcgcga taatgtcggg caatcaggtg cgacaatcta
     4381 tcgattgtat gggaagcccg atgcgccaga gttgtttctg aaacatggca aaggtagcgt
     4441 tgccaatgat gttacagatg agatggtcag actaaactgg ctgacggaat ttatgcctct
     4501 tccgaccatc aagcatttta tccgtactcc tgatgatgca tggttactca ccactgcgat
     4561 ccccgggaaa acagcattcc aggtattaga agaatatcct gattcaggtg aaaatattgt
     4621 tgatgcgctg gcagtgttcc tgcgccggtt gcattcgatt cctgtttgta attgtccttt
     4681 taacagcgat cgcgtatttc gtctcgctca ggcgcaatca cgaatgaata acggtttggt
     4741 tgatgcgagt gattttgatg acgagcgtaa tggctggcct gttgaacaag tctggaaaga
     4801 aatgcataaa cttttgccat tctcaccgga ttcagtcgtc actcatggtg atttctcact
     4861 tgataacctt atttttgacg aggggaaatt aataggttgt attgatgttg gacgagtcgg
     4921 aatcgcagac cgataccagg atcttgccat cctatggaac tgcctcggtg agttttctcc
     4981 ttcattacag aaacggcttt ttcaaaaata tggtattgat aatcctgata tgaataaatt
     5041 gcagtttcat ttgatgctcg atgagttttt ctaagaatta attcatgagc ggatacatat
     5101 ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc
     5161 cacctgaaat tgtaaacgtt aatattttgt taaaattcgc gttaaatttt tgttaaatca
     5221 gctcattttt taaccaatag gccgaaatcg gcaaaatccc ttataaatca aaagaataga
     5281 ccgagatagg gttgagtgtt gttccagttt ggaacaagag tccactatta aagaacgtgg
     5341 actccaacgt caaagggcga aaaaccgtct atcagggcga tggcccacta cgtgaaccat
     5401 caccctaatc aagttttttg gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag
     5461 ggagcccccg atttagagct tgacggggaa agccggcgaa cgtggcgaga aaggaaggga
     5521 agaaagcgaa aggagcgggc gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa
     5581 ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc gtcccattcg cca
//
