LOCUS       sequence-215478-        7851 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Protein expression and purification of ACTG1_Actin.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7851)
  AUTHORS   Teyra J, Kelil A, Jain S, Helmy M, Jajodia R, Hooda Y, Gu J, D'Cruz 
            AA, Nicholson SE, Min J, Sudol M, Kim PM, Bader GD, Sidhu SS
  TITLE     Large-scale survey and database of high affinity ligands for peptide
            recognition modules.
  JOURNAL   Mol Syst Biol. 2020 Dec;16(12):e9310. doi: 10.15252/msb.20199310.
  PUBMED    33438817
REFERENCE   2  (bases 1 to 7851)
  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: "Mol Syst 
            Biol."; date: "2020-12"; volume: "16(12):e9310. doi"; pages: " 
            10.15252/msb.20199310"
COMMENT     Sequence Label: pHH0103_ACTG1_Actin
FEATURES             Location/Qualifiers
     source          1..7851
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(1..6,7..22,23..29)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    37..53
                     /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)."
     primer_bind     42..64
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     CDS             138..155
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             171..824
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     782..804
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             831..854
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             867..884
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     terminator      2046..2093
                     /label=fd terminator
                     /note="central terminator from bacteriophage fd (Otsuka and
                     Kunisawa, 1982)"
     rep_origin      complement(3686..4141)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3823..3844)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4035..4054
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     complement(4183..4205)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4291..4431
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4346..4365
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4446..4463)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4617..5204)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(5375..6166,6167..6235))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     5998..6017
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6236..6340)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6408..6426
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        6449..6526
                     /gene="lacI (mutant)"
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the 
                     promoter boosts expression of the lacI gene about 10-fold."
     CDS             6527..7609
                     /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     complement(6546..6565)
                     /label=LacI-R
                     /note="LacI, reverse primer"
ORIGIN
        1 ttgacaatta atcatcggct cgtataatgt gtggaattgt gagcggataa caatttcaca
       61 caggaaacag ccagtccgtt taggtgtttt cacgagcact tcaccaacaa ggaccataga
      121 ttatgaaaat cgaagaacac catcaccatc accattccag cggtaagctt atgtccccta
      181 tactaggtta ttggaaaatt aagggccttg tgcaacccac tcgacttctt ttggaatatc
      241 ttgaagaaaa atatgaagag catttgtatg agcgcgatga aggtgataaa tggcgaaaca
      301 aaaagtttga attgggtttg gagtttccca atcttcctta ttatattgat ggtgatgtta
      361 aattaacaca gtctatggcc atcatacgtt atatagctga caagcacaac atgttgggtg
      421 gttgtccaaa agagcgtgca gagatttcaa tgcttgaagg agcggttttg gatattagat
      481 acggtgtttc gagaattgca tatagtaaag actttgaaac tctcaaagtt gattttctta
      541 gcaagctacc tgaaatgctg aaaatgttcg aagatcgttt atgtcataaa acatatttaa
      601 atggtgatca tgtaacccat cctgacttca tgttgtatga cgctcttgat gttgttttat
      661 acatggaccc aatgtgcctg gatgcgttcc caaaattagt ttgttttaaa aaacgtattg
      721 aagctatccc acaaattgat aagtacttga aatccagcaa gtatatagca tggcctttgc
      781 agggctggca agccacgttt ggtggtggcg accatcctcc aaaatcggat ctggaagttc
      841 tgttccaggg gcccctgtcc agcggtctgg ttccgcgtgg ttccggtacc gcggcccagc
      901 cggccatgga agaagaaatt gcggccctgg tgatcgataa cggctcaggc atgtgcaaag
      961 caggctttgc tggtgatgac gcgccgcgtg ccgttttccc gtcgattgtc ggccgtccgc
     1021 gccatcaggg tgtgatggtt ggcatgggtc aaaaagatag ttatgtcggc gacgaagcac
     1081 agtccaaacg cggtattctg acgctgaaat acccgattga acatggcatc gtgaccaact
     1141 gggatgacat ggaaaaaatc tggcatcaca ccttttataa tgaactgcgt gtcgccccgg
     1201 aagaacaccc ggtgctgctg acggaagcac cgctgaaccc gaaagctaat cgcgaaaaaa
     1261 tgacccagat tatgtttgaa acgttcaata ccccggcaat gtatgtggca atccaagctg
     1321 ttctgagtct gtacgcttcc ggccgtacca cgggtattgt gatggatagc ggcgacggtg
     1381 tcacgcatac cgtgccgatt tatgaaggtt acgcactgcc gcacgcaatc ctgcgtctgg
     1441 atctggcggg tcgcgatctg acggactatc tgatgaaaat cctgaccgaa cgcggttaca
     1501 gctttaccac gaccgccgaa cgtgaaattg ttcgcgatat caaagaaaaa ctgtgctatg
     1561 tcgcgctgga cttcgaacag gaaatggcca ccgcagctag ctctagttcc ctggaaaaat
     1621 cctacgaact gccggatggc caggttatta ccatcggtaa cgaacgtttt cgctgtccgg
     1681 aagcgctgtt tcaaccgtca ttcctgggca tggaatcgtg cggtattcat gaaacgacct
     1741 tcaacagcat tatgaaatgt gatgtggaca tccgtaaaga tctgtatgcg aataccgttc
     1801 tgtctggcgg tacgaccatg tacccgggca ttgccgaccg catgcagaaa gaaatcacgg
     1861 cgctggcccc gagtaccatg aaaattaaaa ttatcgcacc gccggaacgt aaatatagcg
     1921 tttggattgg cggttctatc ctggcttcac tgtcgacctt tcagcaaatg tggatttcta
     1981 aacaagaata cgatgaaagc ggtccgtcta tcgtccaccg caaatgtttc gcggccgcat
     2041 aataaaccga tacaattaaa ggctcctttt ggagcctttt ttttggagat tttcaacgtg
     2101 aaaaaattat tattcgcaat tcctttagtt gttcctttct attctcactc cgctgaaact
     2161 gttgaaagtt gtttagcaaa accccataca gaaaattcat ttactaacgt ctggaaagac
     2221 gacaaaactt tagatcgtta cgctaactat gagggctgtc tgtggaatgc tacaggcgtt
     2281 gtagtttgta ctggtgacga aactcagtgt ctagctagag tggcggtggc tctggttccg
     2341 gtgattttga ttatgaaaag atggcaaacg ctaataaggg ggctatgacc gaaaatgccg
     2401 atgaaaacgc gctacagtct gacgctaaag gcaaacttga ttctgtcgct actgattacg
     2461 gtgctgctat cgatggtttc attggtgacg tttccggcct tgctaatggt aatggtgcta
     2521 ctggtgattt tgctggctct aattcccaaa tggctcaagt cggtgacggt gataattcac
     2581 ctttaatgaa taatttccgt caatatttac cttccctccc tcaatcggtt gaatgtcgcc
     2641 cttttgtctt tggcgctggt aaaccatatg aattttctat tgattgtgac aaaataaact
     2701 tattccgtgg tgtctttgcg tttcttttat atgttgccac ctttatgtat gtattttcta
     2761 cgtttgctaa catactgcgt aataaggagt cttaatcatg ccagttcttt tggctagcgc
     2821 cgccctatac cttgtctgcc tccccgcgtt gcgtcgcggt gcatggagcc gggccacctc
     2881 gacctgaatg gaagccggcg gcacctcgct aacggattca ccactccaag aattggagcc
     2941 aatcaattct tgcggagaac tgtgaatgcg caaaccaacc cttggcagaa catatccatc
     3001 gcgtccgcca tctccagcag ccgcacgcgg cgcatctcgg gcagcgttgg gtcctggcca
     3061 cgggtgcgca tgatcgtgct cctgtcgttg aggacccggc taggctggcg gggttgcctt
     3121 actggttagc agaatgaatc accgatacgc gagcgaacgt gaagcgactg ctgctgcaaa
     3181 acgtctgcga cctgagcaac aacatgaatg gtcttcggtt tccgtgtttc gtaaagtctg
     3241 gaaacgcgga agtcagcgcc ctgcaccatt atgttccgga tctgcatcgc aggatgctgc
     3301 tggctaccct gtggaacacc tacatctgta ttaacgaagc gctggcattg accctgagtg
     3361 atttttctct ggtcccgccg catccatacc gccagttgtt taccctcaca acgttccagt
     3421 aaccgggcat gttcatcatc agtaacccgt atcgtgagca tcctctctcg tttcatcggt
     3481 atcattaccc ccatgaacag aaatccccct tacacggagg catcagtgac caaacaggaa
     3541 aaaaccgccc ttaacatggc ccgctttatc agaagccaga cattaacgct tctggagaaa
     3601 ctcaacgagc tggacgcgga tgaacaggca gacatctgtg aatcgcttca cgaccacgct
     3661 gatgagcttt accgcaggat ccgcgaaatt gtaagcgtta atattttgtt aaaattcgcg
     3721 ttaaattttt gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg caaaatccct
     3781 tataaatcaa aagaatagac cgagataggg ttgagtgttg ttccagtttg gaacaagagt
     3841 ccactattaa agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat
     3901 ggcccactac gtgaaccatc accctaatca agttttttgg ggtcgaggtg ccgtaaagca
     3961 ctaaatcgga accctaaagg gagcccccga tttagagctt gacggggaaa gccggcgaac
     4021 gtggcgagaa aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta
     4081 gcggtcacgc tgcgcgtaac caccacaccc gccgcgctta atgcgccgct acagggcgcg
     4141 tcgcggatcc tgcctcgcgc gtttcggtga tgacggtgaa aacctctgac acatgcagct
     4201 cccggagacg gtcacagctt gtctgtaagc ggatgccggg agcagacaag cccgtcaggg
     4261 cgcgtcagcg ggtgttggcg ggtgtcgggg cgcagccatg acccagtcac gtagcgatag
     4321 cggagtgtat actggcttaa ctatgcggca tcagagcaga ttgtactgag agtgcaccat
     4381 atgcggtgtg aaataccgca cagatgcgta aggagaaaat accgcatcag gcgctcttcc
     4441 gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct
     4501 cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg
     4561 tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc
     4621 cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga
     4681 aacccgacag gactataaag ataccagggt ttccccctgg aagctccctc gtgcgctctc
     4741 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg
     4801 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc
     4861 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
     4921 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca
     4981 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact
     5041 acggctacac tagaaggaca gtatttggta tctgcgctct gctgaagcca gttaccttcg
     5101 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt
     5161 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct
     5221 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga
     5281 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa
     5341 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac
     5401 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga
     5461 taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc
     5521 cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca
     5581 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta
     5641 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct gcaggcatcg
     5701 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc
     5761 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg
     5821 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt
     5881 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt
     5941 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca acacgggata
     6001 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc
     6061 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac
     6121 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa
     6181 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct
     6241 tcctttttca atattattga agcatttatc agggttattg tctcatgagc ggatacatat
     6301 ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc
     6361 cacctgacgt ctaagaaacc attattatca tgacattaac ctataaaaat aggcgtatca
     6421 cgaggccctt tcgtcttcaa gaattcccga caccatcgaa tggtgcaaaa cctttcgcgg
     6481 tatggcatga tagcgcccgg aagagagtca attcagggtg gtgaatgtga aaccagtaac
     6541 gttatacgat gtcgcagagt atgccggtgt ctcttatcag accgtttccc gcgtggtgaa
     6601 ccaggccagc cacgtttctg cgaaaacgcg ggaaaaagtg gaagcggcga tggcggagct
     6661 gaattacatt cccaaccgcg tggcacaaca actggcgggc aaacagtcgt tgctgattgg
     6721 cgttgccacc tccagtctgg ccctgcacgc gccgtcgcaa attgtcgcgg cgattaaatc
     6781 tcgcgccgat caactgggtg ccagcgtggt ggtgtcgatg gtagaacgaa gcggcgtcga
     6841 agcctgtaaa gcggcggtgc acaatcttct cgcgcaacgc gtcagtgggc tgatcattaa
     6901 ctatccgctg gatgaccagg atgccattgc tgtggaagct gcctgcacta atgttccggc
     6961 gttatttctt gatgtctctg accagacacc catcaacagt attattttct cccatgaaga
     7021 cggtacgcga ctgggcgtgg agcatctggt cgcattgggt caccagcaaa tcgcgctgtt
     7081 agcgggccca ttaagttctg tctcggcgcg tctgcgtctg gctggctggc ataaatatct
     7141 cactcgcaat caaattcagc cgatagcgga acgggaaggc gactggagtg ccatgtccgg
     7201 ttttcaacaa accatgcaaa tgctgaatga gggcatcgtt cccactgcga tgctggttgc
     7261 caacgatcag atggcgctgg gcgcaatgcg cgccattacc gagtccgggc tgcgcgttgg
     7321 tgcggatatc tcggtagtgg gatacgacga taccgaagac agctcatgtt atatcccgcc
     7381 gttaaccacc atcaaacagg attttcgcct gctggggcaa accagcgtgg accgcttgct
     7441 gcaactctct cagggccagg cggtgaaggg caatcagctg ttgcccgtct cactggtgaa
     7501 aagaaaaacc accctggcgc ccaatacgca aaccgcctct ccccgcgcgt tggccgattc
     7561 attaatgcag ctggcacgac aggtttcccg actggaaagc gggcagtgag cgcaacgcaa
     7621 ttaatgtaag ttagctcact cattaggcac aattctcatg tttgacagct tatcatcgac
     7681 tgcacggtgc accaatgctt ctggcgtcag gcagccatcg gaagctgtgg tatggctgtg
     7741 caggtcgtaa atcactgcat aattcgtgtc gctcaaggcg cactcccgtt ctggataatg
     7801 ttttttgcgc cgacatcata acggttctgg caaatattct gaaatgagct g
//
