LOCUS       sequence-227018-        6643 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  SdrG N2 and N3, B1 domains with C-terminal ybbr tag for covalent 
            immobilization.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6643)
  AUTHORS   Milles LF, Unterauer EM, Nicolaus T, Gaub HE
  TITLE     Calcium stabilizes the strongest protein fold.
  JOURNAL   Nat Commun. 2018 Nov 12;9(1):4764. doi: 10.1038/s41467-018-07145-6.
  PUBMED    30420680
REFERENCE   2  (bases 1 to 6643)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Commun."; date: "2018-11-12"; volume: "9(1):4764. doi"; pages: " 
            10.1038/s41467-018-07145-6"
COMMENT     Sequence Label: pET28a-SdrG_N2N3-B1-HIS-ybbr
FEATURES             Location/Qualifiers
     source          1..6643
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     99..118
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        99..117
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    118..142
                     /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             157..179
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             1534..1551
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1558..1581
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     terminator      1690..1737
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1774..2229
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1861..1880)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2071..2092
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(2321..3136)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(2321..3136)
                     /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"
     primer_bind     3044..3063
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     3128..3147
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      3258..3846
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3747..3766
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4000..4017
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4032..4171
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4097..4116)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4257..4279
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4273..4464)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(5273..6355)
                     /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"
     primer_bind     6317..6336
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(6356..6433)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     join(6639..6643,1..15)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 tgtcggcgat ataggcgcca gcaaccgcac ctgtggcgcc ggtgatgccg gccacgatgc
       61 gtccggcgta gaggatcgag atctcgatcc cgcgaaatta atacgactca ctatagggga
      121 attgtgagcg gataacaatt cccctgtaga aataattttg tttaacttta agaaggagat
      181 atacatatgg gtaccgaaca gggtagcaat gtgaaccatc tgattaaagt taccgatcag
      241 agcattaccg aaggctatga tgatagtgat ggtatcatta aagcacacga tgccgaaaac
      301 ctgatttatg atgttacctt tgaggtggat gataaagtga aaagcggtga taccatgacc
      361 gtgaacattg ataaaaacac cgttccgagc gatctgaccg atagctttgc aattccgaaa
      421 atcaaagata acagcggtga aattattgcc accggcacct atgataatac caacaaacaa
      481 atcacctaca ccttcaccga ttacgtggat aaatacgaaa acatcaaagc ccatctgaaa
      541 ctgaccagct atatcgataa aagcaaagtg ccgaacaaca acaccaaact ggatgttgag
      601 tataaaaccg cactgagcag cgtgaataaa accattaccg ttgaatatca gaaaccgaac
      661 gaaaatcgta ccgcaaatct gcagagcatg tttaccaata tcgataccaa aaatcatacc
      721 gtcgaacaga ccatttatat caatccgctg cgttatagcg ccaaagaaac caatgtgaat
      781 attagcggca atggtgatga aggcagcacc attattgatg atagcaccat catcaaagtg
      841 tacaaagtgg gcgataatca gaatctgccg gatagcaatc gcatctatga ttacagcgaa
      901 tatgaagatg tgaccaacga tgattatgca cagctgggta ataataacga cgtgaatatc
      961 aacttcggca atattgatag cccgtatatt atcaaagtga tcagcaaata cgacccgaac
     1021 aaagatgatt acaccaccat tcagcagacc gttaccatgc agaccaccat taatgaatat
     1081 accggtgaat ttcgtaccgc cagctatgat aacacaattg catttagcac cagcagcggt
     1141 cagggtcagg gtgatctgcc tccggaaaaa acctataaaa tcggtgatta tgtgtgggag
     1201 gatgtggata aagatggcat tcagaatacc aacgataacg aaaaaccgct gagcaatgtt
     1261 ctggttaccc tgacctatcc ggatggcacc agcaaaagcg ttcgtaccga tgaagaaggt
     1321 aaatatcagt ttgatggcct gaaaaatggc ctgacctaca aaatcacctt tgaaacaccg
     1381 gaaggttata ccccgaccct gaaacatagc ggcaccaatc cggcactgga tagcgaaggt
     1441 aatagcgttt gggttacaat taatggccag gatgatatga ccattgatag cggcttttat
     1501 cagaccccgg agctcaagct tcctaggtct aggcaccacc accaccacca cggatccctg
     1561 gaagttctgt tccagggtcc ggactctctg gaattcatcg cttctaaact ggcttaactc
     1621 gagtaagatc cggctgctaa caaagcccga aaggaagctg agttggctgc tgccaccgct
     1681 gagcaataac tagcataacc ccttggggcc tctaaacggg tcttgagggg ttttttgctg
     1741 aaaggaggaa ctatatccgg attggcgaat gggacgcgcc ctgtagcggc gcattaagcg
     1801 cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg
     1861 ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc
     1921 taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa
     1981 aacttgatta gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc
     2041 ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac
     2101 tcaaccctat ctcggtctat tcttttgatt tataagggat tttgccgatt tcggcctatt
     2161 ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa atattaacgc
     2221 ttacaattta ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt
     2281 ctaaatacat tcaaatatgt atccgctcat gaattaattc ttagaaaaac tcatcgagca
     2341 tcaaatgaaa ctgcaattta ttcatatcag gattatcaat accatatttt tgaaaaagcc
     2401 gtttctgtaa tgaaggagaa aactcaccga ggcagttcca taggatggca agatcctggt
     2461 atcggtctgc gattccgact cgtccaacat caatacaacc tattaatttc ccctcgtcaa
     2521 aaataaggtt atcaagtgag aaatcaccat gagtgacgac tgaatccggt gagaatggca
     2581 aaagtttatg catttctttc cagacttgtt caacaggcca gccattacgc tcgtcatcaa
     2641 aatcactcgc atcaaccaaa ccgttattca ttcgtgattg cgcctgagcg agacgaaata
     2701 cgcgatcgct gttaaaagga caattacaaa caggaatcga atgcaaccgg cgcaggaaca
     2761 ctgccagcgc atcaacaata ttttcacctg aatcaggata ttcttctaat acctggaatg
     2821 ctgttttccc ggggatcgca gtggtgagta accatgcatc atcaggagta cggataaaat
     2881 gcttgatggt cggaagaggc ataaattccg tcagccagtt tagtctgacc atctcatctg
     2941 taacatcatt ggcaacgcta cctttgccat gtttcagaaa caactctggc gcatcgggct
     3001 tcccatacaa tcgatagatt gtcgcacctg attgcccgac attatcgcga gcccatttat
     3061 acccatataa atcagcatcc atgttggaat ttaatcgcgg cctagagcaa gacgtttccc
     3121 gttgaatatg gctcataaca ccccttgtat tactgtttat gtaagcagac agttttattg
     3181 ttcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa
     3241 aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca
     3301 aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt
     3361 ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgtccttct agtgtagccg
     3421 tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc
     3481 ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga
     3541 cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc
     3601 agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc
     3661 gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca
     3721 ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg
     3781 tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta
     3841 tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct
     3901 cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag
     3961 tgagctgata ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa
     4021 gcggaagagc gcctgatgcg gtattttctc cttacgcatc tgtgcggtat ttcacaccgc
     4081 aatggtgcac tctcagtaca atctgctctg atgccgcata gttaagccag tatacactcc
     4141 gctatcgcta cgtgactggg tcatggctgc gccccgacac ccgccaacac ccgctgacgc
     4201 gccctgacgg gcttgtctgc tcccggcatc cgcttacaga caagctgtga ccgtctccgg
     4261 gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgaggc agctgcggta
     4321 aagctcatca gcgtggtcgt gaagcgattc acagatgtct gcctgttcat ccgcgtccag
     4381 ctcgttgagt ttctccagaa gcgttaatgt ctggcttctg ataaagcggg ccatgttaag
     4441 ggcggttttt tcctgtttgg tcactgatgc ctccgtgtaa gggggatttc tgttcatggg
     4501 ggtaatgata ccgatgaaac gagagaggat gctcacgata cgggttactg atgatgaaca
     4561 tgcccggtta ctggaacgtt gtgagggtaa acaactggcg gtatggatgc ggcgggacca
     4621 gagaaaaatc actcagggtc aatgccagcg cttcgttaat acagatgtag gtgttccaca
     4681 gggtagccag cagcatcctg cgatgcagat ccggaacata atggtgcagg gcgctgactt
     4741 ccgcgtttcc agactttacg aaacacggaa accgaagacc attcatgttg ttgctcaggt
     4801 cgcagacgtt ttgcagcagc agtcgcttca cgttcgctcg cgtatcggtg attcattctg
     4861 ctaaccagta aggcaacccc gccagcctag ccgggtcctc aacgacagga gcacgatcat
     4921 gcgcacccgt ggggccgcca tgccggcgat aatggcctgc ttctcgccga aacgtttggt
     4981 ggcgggacca gtgacgaagg cttgagcgag ggcgtgcaag attccgaata ccgcaagcga
     5041 caggccgatc atcgtcgcgc tccagcgaaa gcggtcctcg ccgaaaatga cccagagcgc
     5101 tgccggcacc tgtcctacga gttgcatgat aaagaagaca gtcataagtg cggcgacgat
     5161 agtcatgccc cgcgcccacc ggaaggagct gactgggttg aaggctctca agggcatcgg
     5221 tcgagatccc ggtgcctaat gagtgagcta acttacatta attgcgttgc gctcactgcc
     5281 cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg
     5341 gagaggcggt ttgcgtattg ggcgccaggg tggtttttct tttcaccagt gagacgggca
     5401 acagctgatt gcccttcacc gcctggccct gagagagttg cagcaagcgg tccacgctgg
     5461 tttgccccag caggcgaaaa tcctgtttga tggtggttaa cggcgggata taacatgagc
     5521 tgtcttcggt atcgtcgtat cccactaccg agatatccgc accaacgcgc agcccggact
     5581 cggtaatggc gcgcattgcg cccagcgcca tctgatcgtt ggcaaccagc atcgcagtgg
     5641 gaacgatgcc ctcattcagc atttgcatgg tttgttgaaa accggacatg gcactccagt
     5701 cgccttcccg ttccgctatc ggctgaattt gattgcgagt gagatattta tgccagccag
     5761 ccagacgcag acgcgccgag acagaactta atgggcccgc taacagcgcg atttgctggt
     5821 gacccaatgc gaccagatgc tccacgccca gtcgcgtacc gtcttcatgg gagaaaataa
     5881 tactgttgat gggtgtctgg tcagagacat caagaaataa cgccggaaca ttagtgcagg
     5941 cagcttccac agcaatggca tcctggtcat ccagcggata gttaatgatc agcccactga
     6001 cgcgttgcgc gagaagattg tgcaccgccg ctttacaggc ttcgacgccg cttcgttcta
     6061 ccatcgacac caccacgctg gcacccagtt gatcggcgcg agatttaatc gccgcgacaa
     6121 tttgcgacgg cgcgtgcagg gccagactgg aggtggcaac gccaatcagc aacgactgtt
     6181 tgcccgccag ttgttgtgcc acgcggttgg gaatgtaatt cagctccgcc atcgccgctt
     6241 ccactttttc ccgcgttttc gcagaaacgt ggctggcctg gttcaccacg cgggaaacgg
     6301 tctgataaga gacaccggca tactctgcga catcgtataa cgttactggt ttcacattca
     6361 ccaccctgaa ttgactctct tccgggcgct atcatgccat accgcgaaag gttttgcgcc
     6421 attcgatggt gtccgggatc tcgacgctct cccttatgcg actcctgcat taggaagcag
     6481 cccagtagta ggttgaggcc gttgagcacc gccgccgcaa ggaatggtgc atgcaaggag
     6541 atggcgccca acagtccccc ggccacgggg cctgccacca tacccacgcc gaaacaagcg
     6601 ctcatgagcc cgaagtggcg agcccgatct tccccatcgg tga
//
