LOCUS       A_pET-29b(+)_His        6461 bp DNA     circular SYN 05-MAY-2021
DEFINITION  clone verified 11.02.21.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6461)
  AUTHORS   kelvinlau
  TITLE     Direct Submission
  JOURNAL   Exported Mar 13, 2024 from SnapGene 7.1.2
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6461
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        3..21
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
                     /note="color: #ffffff; direction: RIGHT"
     protein_bind    22..46
                     /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)."
                     /note="color: #31849b"
     RBS             61..83
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
                     /note="color: #a6acb3"
     mat_peptide     94..105
                     /label=tufA enhancer peptide
                     /note="Varma 2019 Nature Comm"
                     /note="color: #cc99b2; direction: RIGHT"
     CDS             106..135
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /note="color: #cc99b2"
                     /translation="HHHHHHHHHH"
     CDS             154..309
                     /codon_start=1
                     /gene="Bacillus circulans chiA"
                     /product="chitin binding domain from chitinase A1 (Watanabe
                     et al., 1994)"
                     /label=CBD
                     /note="color: #cc99b2"
                     /translation="TTNPGVSAWQVNTAYTAGQLVTYNGKTYKCLQPHTSLAGWEPSNV
                     PALWQLQ"
     CDS             316..336
                     /codon_start=1
                     /product="DYDIPTT"
                     /label=TEV cleavage enhancer element
                     /note="10.1016/j.cell.2006.05.046"
                     /note="color: #ff00ff"
                     /translation="DYDIPTT"
     CDS             340..363
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /note="color: #cc99b2
                     Cleavage site after base 357"
                     /translation="LEVLFQGP"
     CDS             370..1311
                     /codon_start=1
                     /label=PNGaseF 41-354
                     /note="P21163 (PNGF_ELIMR)"
                     /note="color: #993366"
                     /translation="APADNTVNIKTFDKVKNAFGDGLSQSAEGTFTFPADVTTVKTIKM
                     FIKNECPNKTCDEWDRYANVYVKNKTTGEWYEIGRFITPYWVGTEKLPRGLEIDVTDFK
                     SLLSGNTELKIYTETWLAKGREYSVDFDIVYGTPDYKYSAVVPVIQYNKSSIDGVPYGK
                     AHTLGLKKNIQLPTNTEKAYLRTTISGWGHAKPYDAGSRGCAEWCFRTHTIAINNANTF
                     QHQLGALGCSANPINNQSPGNWAPDRAGWCPGMAVPTRIDVLNNSLTGSTFSYEYKFQS
                     WTNNGTNGDAFYAISSFVIAKSNTPISAPVVTN"
     CDS             1324..1341
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /note="color: #cc99b2"
                     /translation="HHHHHH"
     terminator      1408..1455
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
                     /note="color: #ffffff"
     rep_origin      1492..1947
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
                     /note="color: #ffff00"
     CDS             complement(2039..2854)
                     /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"
                     /note="color: #ccffcc"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     rep_origin      2976..3564
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
                     /note="color: #ffff00"
     misc_feature    3750..3889
                     /label=bom
                     /note="basis of mobility region from pBR322"
                     /note="color: #a6acb3"
     CDS             complement(3991..4182)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /note="color: #993366"
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4991..6073)
                     /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)."
                     /note="color: #993366"
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     promoter        complement(6074..6151)
                     /gene="lacI"
                     /label=lacI promoter
                     /note="color: #ffffff; direction: LEFT"
ORIGIN
        1 attaatacga ctcactatag gggaattgtg agcggataac aattcccctc tagaaataat
       61 tttgtttaac tttaagaagg agatatacat atgtctaaag aaaaacacca tcatcatcat
      121 catcaccatc accatggatc tgccaactca ggcaccacga atccaggcgt gtcagcctgg
      181 caagtgaaca ccgcctacac agcagggcag ttggtcacgt ataacggcaa aacgtataaa
      241 tgccttcaac ctcacacaag tcttgccggt tgggagccat ccaacgtacc cgccttatgg
      301 caactgcaat ctggtgacta cgatattccg actaccggat tggaggtatt gttccaaggt
      361 ccaggatccg cgccggccga taacacggtc aatattaaga cgttcgacaa agtcaagaat
      421 gctttcggag acgggttgag ccaaagtgcc gaagggacgt tcacctttcc ggcggacgta
      481 acaacggtca aaactattaa aatgttcatc aaaaatgagt gccctaataa aacatgcgac
      541 gaatgggatc gctatgctaa tgtatatgtc aagaacaaga ccaccggtga atggtatgaa
      601 atcggacgtt tcattacacc ttactgggtc ggcaccgaga aattgccccg tggactggag
      661 attgatgtta cggactttaa gtcattactt agtggaaata cagagcttaa aatctacacc
      721 gagacctggt tggccaaggg ccgcgaatat agtgtcgatt ttgacatcgt gtatggcacc
      781 ccggactata agtacagtgc cgtggtcccc gtcattcaat acaacaaatc ttccatcgac
      841 ggggtcccgt atggtaaagc gcatacatta gggctgaaga agaacatcca attgccaaca
      901 aacaccgaga aggcatattt gcgcaccact atcagcggct ggggccatgc taaaccatac
      961 gatgctggct cgcgtggttg tgcggagtgg tgttttcgca cccatacaat tgccatcaac
     1021 aacgctaaca cgttccaaca tcagttagga gctttagggt gtagtgccaa tcctatcaac
     1081 aaccaatcac ctgggaattg ggcgccagat cgcgcaggtt ggtgtcctgg aatggcagtt
     1141 cctacgcgta ttgacgttct gaataactcc ctgactggat ctactttctc gtatgagtac
     1201 aagttccaat cgtggacgaa caacgggacc aatggcgacg ccttctacgc aatttcatca
     1261 tttgtaatcg caaaatctaa tactcccatc tcggcacctg tagttacgaa ttaatagctc
     1321 gagcaccacc accaccacca ctgagatccg gctgctaaca aagcccgaaa ggaagctgag
     1381 ttggctgctg ccaccgctga gcaataacta gcataacccc ttggggcctc taaacgggtc
     1441 ttgaggggtt ttttgctgaa aggaggaact atatccggat tggcgaatgg gacgcgccct
     1501 gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg
     1561 ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg
     1621 gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac
     1681 ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtagtggg ccatcgccct
     1741 gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt
     1801 tccaaactgg aacaacactc aaccctatct cggtctattc ttttgattta taagggattt
     1861 tgccgatttc ggcctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt
     1921 ttaacaaaat attaacgctt acaatttagg tggcactttt cggggaaatg tgcgcggaac
     1981 ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga attaattctt
     2041 agaaaaactc atcgagcatc aaatgaaact gcaatttatt catatcagga ttatcaatac
     2101 catatttttg aaaaagccgt ttctgtaatg aaggagaaaa ctcaccgagg cagttccata
     2161 ggatggcaag atcctggtat cggtctgcga ttccgactcg tccaacatca atacaaccta
     2221 ttaatttccc ctcgtcaaaa ataaggttat caagtgagaa atcaccatga gtgacgactg
     2281 aatccggtga gaatggcaaa agtttatgca tttctttcca gacttgttca acaggccagc
     2341 cattacgctc gtcatcaaaa tcactcgcat caaccaaacc gttattcatt cgtgattgcg
     2401 cctgagcgag acgaaatacg cgatcgctgt taaaaggaca attacaaaca ggaatcgaat
     2461 gcaaccggcg caggaacact gccagcgcat caacaatatt ttcacctgaa tcaggatatt
     2521 cttctaatac ctggaatgct gttttcccgg ggatcgcagt ggtgagtaac catgcatcat
     2581 caggagtacg gataaaatgc ttgatggtcg gaagaggcat aaattccgtc agccagttta
     2641 gtctgaccat ctcatctgta acatcattgg caacgctacc tttgccatgt ttcagaaaca
     2701 actctggcgc atcgggcttc ccatacaatc gatagattgt cgcacctgat tgcccgacat
     2761 tatcgcgagc ccatttatac ccatataaat cagcatccat gttggaattt aatcgcggcc
     2821 tagagcaaga cgtttcccgt tgaatatggc tcataacacc ccttgtatta ctgtttatgt
     2881 aagcagacag ttttattgtt catgaccaaa atcccttaac gtgagttttc gttccactga
     2941 gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta
     3001 atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa
     3061 gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact
     3121 gtccttctag tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca
     3181 tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt
     3241 accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg
     3301 ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag
     3361 cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta
     3421 agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat
     3481 ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg
     3541 tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc
     3601 ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac
     3661 cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc
     3721 gagtcagtga gcgaggaagc ggaagagcgc ctgatgcggt attttctcct tacgcatctg
     3781 tgcggtattt cacaccgcaa tggtgcactc tcagtacaat ctgctctgat gccgcatagt
     3841 taagccagta tacactccgc tatcgctacg tgactgggtc atggctgcgc cccgacaccc
     3901 gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg cttacagaca
     3961 agctgtgacc gtctccggga gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg
     4021 cgcgaggcag ctgcggtaaa gctcatcagc gtggtcgtga agcgattcac agatgtctgc
     4081 ctgttcatcc gcgtccagct cgttgagttt ctccagaagc gttaatgtct ggcttctgat
     4141 aaagcgggcc atgttaaggg cggttttttc ctgtttggtc actgatgcct ccgtgtaagg
     4201 gggatttctg ttcatggggg taatgatacc gatgaaacga gagaggatgc tcacgatacg
     4261 ggttactgat gatgaacatg cccggttact ggaacgttgt gagggtaaac aactggcggt
     4321 atggatgcgg cgggaccaga gaaaaatcac tcagggtcaa tgccagcgct tcgttaatac
     4381 agatgtaggt gttccacagg gtagccagca gcatcctgcg atgcagatcc ggaacataat
     4441 ggtgcagggc gctgacttcc gcgtttccag actttacgaa acacggaaac cgaagaccat
     4501 tcatgttgtt gctcaggtcg cagacgtttt gcagcagcag tcgcttcacg ttcgctcgcg
     4561 tatcggtgat tcattctgct aaccagtaag gcaaccccgc cagcctagcc gggtcctcaa
     4621 cgacaggagc acgatcatgc gcacccgtgg ggccgccatg ccggcgataa tggcctgctt
     4681 ctcgccgaaa cgtttggtgg cgggaccagt gacgaaggct tgagcgaggg cgtgcaagat
     4741 tccgaatacc gcaagcgaca ggccgatcat cgtcgcgctc cagcgaaagc ggtcctcgcc
     4801 gaaaatgacc cagagcgctg ccggcacctg tcctacgagt tgcatgataa agaagacagt
     4861 cataagtgcg gcgacgatag tcatgccccg cgcccaccgg aaggagctga ctgggttgaa
     4921 ggctctcaag ggcatcggtc gagatcccgg tgcctaatga gtgagctaac ttacattaat
     4981 tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg
     5041 aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgccagggtg gtttttcttt
     5101 tcaccagtga gacgggcaac agctgattgc ccttcaccgc ctggccctga gagagttgca
     5161 gcaagcggtc cacgctggtt tgccccagca ggcgaaaatc ctgtttgatg gtggttaacg
     5221 gcgggatata acatgagctg tcttcggtat cgtcgtatcc cactaccgag atgtccgcac
     5281 caacgcgcag cccggactcg gtaatggcgc gcattgcgcc cagcgccatc tgatcgttgg
     5341 caaccagcat cgcagtggga acgatgccct cattcagcat ttgcatggtt tgttgaaaac
     5401 cggacatggc actccagtcg ccttcccgtt ccgctatcgg ctgaatttga ttgcgagtga
     5461 gatatttatg ccagccagcc agacgcagac gcgccgagac agaacttaat gggcccgcta
     5521 acagcgcgat ttgctggtga cccaatgcga ccagatgctc cacgcccagt cgcgtaccgt
     5581 cttcatggga gaaaataata ctgttgatgg gtgtctggtc agagacatca agaaataacg
     5641 ccggaacatt agtgcaggca gcttccacag caatggcatc ctggtcatcc agcggatagt
     5701 taatgatcag cccactgacg cgttgcgcga gaagattgtg caccgccgct ttacaggctt
     5761 cgacgccgct tcgttctacc atcgacacca ccacgctggc acccagttga tcggcgcgag
     5821 atttaatcgc cgcgacaatt tgcgacggcg cgtgcagggc cagactggag gtggcaacgc
     5881 caatcagcaa cgactgtttg cccgccagtt gttgtgccac gcggttggga atgtaattca
     5941 gctccgccat cgccgcttcc actttttccc gcgttttcgc agaaacgtgg ctggcctggt
     6001 tcaccacgcg ggaaacggtc tgataagaga caccggcata ctctgcgaca tcgtataacg
     6061 ttactggttt cacattcacc accctgaatt gactctcttc cgggcgctat catgccatac
     6121 cgcgaaaggt tttgcgccat tcgatggtgt ccgggatctc gacgctctcc cttatgcgac
     6181 tcctgcatta ggaagcagcc cagtagtagg ttgaggccgt tgagcaccgc cgccgcaagg
     6241 aatggtgcat gcaaggagat ggcgcccaac agtcccccgg ccacggggcc tgccaccata
     6301 cccacgccga aacaagcgct catgagcccg aagtggcgag cccgatcttc cccatcggtg
     6361 atgtcggcga tataggcgcc agcaaccgca cctgtggcgc cggtgatgcc ggccacgatg
     6421 cgtccggcgt agaggatcga gatcgatctc gatcccgcga a
//
