LOCUS       sequence-181446-        6502 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  CohIII-CBM handle with C-Terminal Sortase Recognition Sequence 
            LPETGG for Posttranslational Coupling to a N-terminal GGG.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6502)
  AUTHORS   Ott W, Jobst MA, Bauer MS, Durner E, Milles LF, Nash MA, Gaub HE
  TITLE     Elastin-like Polypeptide Linkers for Single-Molecule Force 
            Spectroscopy.
  JOURNAL   ACS Nano. 2017 Jun 12. doi: 10.1021/acsnano.7b02694.
  PUBMED    28591514
REFERENCE   2  (bases 1 to 6502)
  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: "ACS Nano. 
            2017 Jun 12. doi: 10.1021/acsnano.7b02694."
COMMENT     Sequence Label: pET28a-CohIII-CBM-HIS-LPETGG
FEATURES             Location/Qualifiers
     source          1..6502
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     971..990
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(1010..1087)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     1293..1312
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     1396..1415
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1396..1414
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1415..1439
                     /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             1454..1476
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             2705..2722
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             2729..2752
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     primer_bind     complement(2832..2850)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2846..2893
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2930..3385
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3017..3036)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3227..3248
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(3477..4292)
                     /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(3477..4292)
                     /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     4200..4219
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     4284..4303
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      4414..5002
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4903..4922
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5156..5173
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    5188..5327
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(5253..5272)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5413..5435
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(5429..5620)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(join(6429..6502,1..1009))
                     /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"
ORIGIN
        1 cggtttgcgt attgggcgcc agggtggttt ttcttttcac cagtgagacg ggcaacagct
       61 gattgccctt caccgcctgg ccctgagaga gttgcagcaa gcggtccacg ctggtttgcc
      121 ccagcaggcg aaaatcctgt ttgatggtgg ttaacggcgg gatataacat gagctgtctt
      181 cggtatcgtc gtatcccact accgagatat ccgcaccaac gcgcagcccg gactcggtaa
      241 tggcgcgcat tgcgcccagc gccatctgat cgttggcaac cagcatcgca gtgggaacga
      301 tgccctcatt cagcatttgc atggtttgtt gaaaaccgga catggcactc cagtcgcctt
      361 cccgttccgc tatcggctga atttgattgc gagtgagata tttatgccag ccagccagac
      421 gcagacgcgc cgagacagaa cttaatgggc ccgctaacag cgcgatttgc tggtgaccca
      481 atgcgaccag atgctccacg cccagtcgcg taccgtcttc atgggagaaa ataatactgt
      541 tgatgggtgt ctggtcagag acatcaagaa ataacgccgg aacattagtg caggcagctt
      601 ccacagcaat ggcatcctgg tcatccagcg gatagttaat gatcagccca ctgacgcgtt
      661 gcgcgagaag attgtgcacc gccgctttac aggcttcgac gccgcttcgt tctaccatcg
      721 acaccaccac gctggcaccc agttgatcgg cgcgagattt aatcgccgcg acaatttgcg
      781 acggcgcgtg cagggccaga ctggaggtgg caacgccaat cagcaacgac tgtttgcccg
      841 ccagttgttg tgccacgcgg ttgggaatgt aattcagctc cgccatcgcc gcttccactt
      901 tttcccgcgt tttcgcagaa acgtggctgg cctggttcac cacgcgggaa acggtctgat
      961 aagagacacc ggcatactct gcgacatcgt ataacgttac tggtttcaca ttcaccaccc
     1021 tgaattgact ctcttccggg cgctatcatg ccataccgcg aaaggttttg cgccattcga
     1081 tggtgtccgg gatctcgacg ctctccctta tgcgactcct gcattaggaa gcagcccagt
     1141 agtaggttga ggccgttgag caccgccgcc gcaaggaatg gtgcatgcaa ggagatggcg
     1201 cccaacagtc ccccggccac ggggcctgcc accataccca cgccgaaaca agcgctcatg
     1261 agcccgaagt ggcgagcccg atcttcccca tcggtgatgt cggcgatata ggcgccagca
     1321 accgcacctg tggcgccggt gatgccggcc acgatgcgtc cggcgtagag gatcgagatc
     1381 tcgatcccgc gaaattaata cgactcacta taggggaatt gtgagcggat aacaattccc
     1441 ctgtagaaat aattttgttt aactttaaga aggagatata catatggcgc tcacagacag
     1501 aggaatgact tacgatctcg atcctaagga tggatcatct gctgctacaa agccagttct
     1561 cgaagtaact aagaaggttt tcgacacagc tgctgatgct gctggccaga ctgttacagt
     1621 tgaattcaag gttagcggtg cagaaggcaa gtacgctaca actggttatc acatctactg
     1681 ggatgagaga cttgaagttg ttgcaaccaa gacaggtgca tacgctaaga agggcgcagc
     1741 tcttgaagat tcaagcttag ctaaggctga gaacaacggt aacggtgttt tcgttgcttc
     1801 aggtgctgat gatgattttg gtgctgacgg tgtaatgtgg actgttgaac tcaaggttcc
     1861 agcagacgct aaggctggcg atgtatatcc aatcgacgtt gcttatcagt gggatccttc
     1921 aaagggtgac cttttcacag ataacaagga ctcagcacag ggcaagctga tgcaggctta
     1981 cttcttcaca cagggtatca agtcatcttc taatccttca acagatgagt acctcgtaaa
     2041 ggctaacgct acatatgctg atggttacat tgcaatcaag gctggtgagc caggcagtgt
     2101 agtaccatca acacagcctg taacaacacc acctgcaaca acaaaaccac ctgcaacaac
     2161 aataccgccg tcagatgatc cgaatgccat ggccaataca ccggtatcag gcaatttgaa
     2221 ggttgaattc tacaacagca atccttcaga tactactaac tcaatcaatc ctcagttcaa
     2281 ggttactaat accggaagca gtgcaattga tttgtccaaa ctcacattga gatattatta
     2341 tacagtagac ggtcagaaag atcagacctt ctggagtgac catgctgcaa taatcggcag
     2401 taacggcagc tacaacggaa ttacttcaaa tgtaaaagga acatttgtaa aaatgagttc
     2461 ctcaacaaat aacgcagaca cctaccttga aataagcttt acaggcggaa ctcttgaacc
     2521 gggtgcacat gttcagatac aaggtagatt tgcaaagaat gactggagta actatacaca
     2581 gtcaaatgac tactcattca agtctgcttc acagtttgtt gaatgggatc aggtaacagc
     2641 atacttgaac ggtgttcttg tatggggtaa agaacccggt gagctcaagc ttcctaggtc
     2701 taggcaccac caccaccacc acggatccct ggaagttctg ttccagggtc cgctgccgga
     2761 aaccggcggc taactcgagt aagatccggc tgctaacaaa gcccgaaagg aagctgagtt
     2821 ggctgctgcc accgctgagc aataactagc ataacccctt ggggcctcta aacgggtctt
     2881 gaggggtttt ttgctgaaag gaggaactat atccggattg gcgaatggga cgcgccctgt
     2941 agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc
     3001 agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc
     3061 tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg
     3121 cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc atcgccctga
     3181 tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc
     3241 caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata agggattttg
     3301 ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt
     3361 aacaaaatat taacgcttac aatttaggtg gcacttttcg gggaaatgtg cgcggaaccc
     3421 ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaat taattcttag
     3481 aaaaactcat cgagcatcaa atgaaactgc aatttattca tatcaggatt atcaatacca
     3541 tatttttgaa aaagccgttt ctgtaatgaa ggagaaaact caccgaggca gttccatagg
     3601 atggcaagat cctggtatcg gtctgcgatt ccgactcgtc caacatcaat acaacctatt
     3661 aatttcccct cgtcaaaaat aaggttatca agtgagaaat caccatgagt gacgactgaa
     3721 tccggtgaga atggcaaaag tttatgcatt tctttccaga cttgttcaac aggccagcca
     3781 ttacgctcgt catcaaaatc actcgcatca accaaaccgt tattcattcg tgattgcgcc
     3841 tgagcgagac gaaatacgcg atcgctgtta aaaggacaat tacaaacagg aatcgaatgc
     3901 aaccggcgca ggaacactgc cagcgcatca acaatatttt cacctgaatc aggatattct
     3961 tctaatacct ggaatgctgt tttcccgggg atcgcagtgg tgagtaacca tgcatcatca
     4021 ggagtacgga taaaatgctt gatggtcgga agaggcataa attccgtcag ccagtttagt
     4081 ctgaccatct catctgtaac atcattggca acgctacctt tgccatgttt cagaaacaac
     4141 tctggcgcat cgggcttccc atacaatcga tagattgtcg cacctgattg cccgacatta
     4201 tcgcgagccc atttataccc atataaatca gcatccatgt tggaatttaa tcgcggccta
     4261 gagcaagacg tttcccgttg aatatggctc ataacacccc ttgtattact gtttatgtaa
     4321 gcagacagtt ttattgttca tgaccaaaat cccttaacgt gagttttcgt tccactgagc
     4381 gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat
     4441 ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga
     4501 gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt
     4561 tcttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata
     4621 cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac
     4681 cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg
     4741 ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg
     4801 tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag
     4861 cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct
     4921 ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc
     4981 aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt
     5041 ttgctggcct tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg
     5101 tattaccgcc tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga
     5161 gtcagtgagc gaggaagcgg aagagcgcct gatgcggtat tttctcctta cgcatctgtg
     5221 cggtatttca caccgcaatg gtgcactctc agtacaatct gctctgatgc cgcatagtta
     5281 agccagtata cactccgcta tcgctacgtg actgggtcat ggctgcgccc cgacacccgc
     5341 caacacccgc tgacgcgccc tgacgggctt gtctgctccc ggcatccgct tacagacaag
     5401 ctgtgaccgt ctccgggagc tgcatgtgtc agaggttttc accgtcatca ccgaaacgcg
     5461 cgaggcagct gcggtaaagc tcatcagcgt ggtcgtgaag cgattcacag atgtctgcct
     5521 gttcatccgc gtccagctcg ttgagtttct ccagaagcgt taatgtctgg cttctgataa
     5581 agcgggccat gttaagggcg gttttttcct gtttggtcac tgatgcctcc gtgtaagggg
     5641 gatttctgtt catgggggta atgataccga tgaaacgaga gaggatgctc acgatacggg
     5701 ttactgatga tgaacatgcc cggttactgg aacgttgtga gggtaaacaa ctggcggtat
     5761 ggatgcggcg ggaccagaga aaaatcactc agggtcaatg ccagcgcttc gttaatacag
     5821 atgtaggtgt tccacagggt agccagcagc atcctgcgat gcagatccgg aacataatgg
     5881 tgcagggcgc tgacttccgc gtttccagac tttacgaaac acggaaaccg aagaccattc
     5941 atgttgttgc tcaggtcgca gacgttttgc agcagcagtc gcttcacgtt cgctcgcgta
     6001 tcggtgattc attctgctaa ccagtaaggc aaccccgcca gcctagccgg gtcctcaacg
     6061 acaggagcac gatcatgcgc acccgtgggg ccgccatgcc ggcgataatg gcctgcttct
     6121 cgccgaaacg tttggtggcg ggaccagtga cgaaggcttg agcgagggcg tgcaagattc
     6181 cgaataccgc aagcgacagg ccgatcatcg tcgcgctcca gcgaaagcgg tcctcgccga
     6241 aaatgaccca gagcgctgcc ggcacctgtc ctacgagttg catgataaag aagacagtca
     6301 taagtgcggc gacgatagtc atgccccgcg cccaccggaa ggagctgact gggttgaagg
     6361 ctctcaaggg catcggtcga gatcccggtg cctaatgagt gagctaactt acattaattg
     6421 cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa
     6481 tcggccaacg cgcggggaga gg
//
