LOCUS       sequence-243752-        7701 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  To transform BL21-Gold(DE3) and induced by IPTG for His-proteinG-Tnp
            (hyperactive mutant) fusion protein..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7701)
  AUTHORS   Zhao Lab
  TITLE     Bacterial expression plasmid for production of recombinant 
            His-proteinG-Tnp fusion protein.
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7701)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pET15bHis6-pG-Tnp
FEATURES             Location/Qualifiers
     source          1..7701
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             19..36
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             46..63
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     primer_bind     complement(2121..2139)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2135..2182
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(join(2357..2362,2363..2379,2380..2385))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     primer_bind     complement(2412..2430)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2498..2602
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2603..2671,2672..3463)
                     /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     complement(2821..2840)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3634..4222
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4123..4142
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4376..4393
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4408..4547
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4473..4492)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4633..4655
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4649..4840)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(6152..7234)
                     /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     7196..7215
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(7235..7312)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     7518..7537
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     7621..7640
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        7621..7639
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    7640..7664
                     /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             7679..7701
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
ORIGIN
        1 tataccatgg gcagcagcca tcatcatcat catcacagca gcggcctggt gccgcgcggc
       61 agccatttga agggtgaaac gaccacagaa gcagtagacg cggccactgc cgagaaggtg
      121 tttaaacaat atgcgaacga caacggagtc gatggggaat ggacttatga cgatgcaacc
      181 aaaactttca ctgttacgga gaagcccgaa gtaattgacg ccagcgaact tacccccgcg
      241 gtaacgacgt ataaacttgt aattaatggt aagacgctga agggggagac caccactgag
      301 gccgtagatg cagctacagc cgagaaggtc tttaaacagt atgccaatga taatggtgtc
      361 gacggtgagt ggacgtacga tgatgcaact aagacattca ccgtaacaga aaagccggaa
      421 gttattgacg cgtcggaact tactcctgcg gtgaccactt acaaactggt aatcaacggc
      481 aagacgctta agggagaaac aacgactaaa gccgtcgacg ccgaaacggc tgagaaagct
      541 ttcaagcaat acgccaatga taatggggta gacggggttt ggacgtatga tgacgccacc
      601 aagactttta cagtcacgga aatgggtggt ggaatgataa cttctgctct tcatcgtgcg
      661 gccgactggg ctaaatctgt gttctcttcg gcggcgctgg gtgatcctcg ccgtactgcc
      721 cgcttggtta acgtcgccgc ccaattggca aaatattctg gtaaatcaat aaccatctca
      781 tcagagggta gtaaagccgc gcaggaaggc gcttaccgat ttatccgcaa tcccaacgtt
      841 tctgccgagg cgatcagaaa ggctggcgcc atgcaaacag tcaagttggc tcaggagttt
      901 cccgaactgc tggccattga ggacaccacc tctttgagtt atcgccacca ggtcgccgaa
      961 gagcttggca agctgggctc tattcaggat aaatcccgcg gatggtgggt tcactccgtt
     1021 ctcttgctcg aggccaccac attccgcacc gtaggattac tgcatcagga gtggtggatg
     1081 cgcccggatg accctgccga tgcggatgaa aaggagagtg gcaaatggct ggcagcggcc
     1141 gcaactagcc ggttacgcat gggcagcatg atgagcaacg tgattgcggt ctgtgaccgc
     1201 gaagccgata ttcatgctta tctgcaggac aaactggcgc ataacgagcg cttcgtggtg
     1261 cgctccaagc acccacgcaa ggacgtagag tctgggttgt atctgtacga ccatctgaag
     1321 aaccaaccgg agttgggtgg ctatcagatc agcattccgc aaaagggcgt ggtggataaa
     1381 cgcggtaaac gtaaaaatcg accagcccgc aaggcgagct tgagcctgcg cagtgggcgc
     1441 atcacgctaa aacaggggaa tatcacgctc aacgcggtgc tggccgagga gattaacccg
     1501 cccaagggtg agaccccgtt gaaatggttg ttgctgacca gcgaaccggt cgagtcgcta
     1561 gcccaagcct tgcgcgtcat cgacatttat acccatcgct ggcggatcga ggagttccat
     1621 aaggcatgga aaaccggagc aggagccgag aggcaacgca tggaggagcc ggataatctg
     1681 gagcggatgg tctcgatcct ctcgtttgtt gcggtcaggc tgttacagct cagagaaagc
     1741 ttcacgccgc cgcaagcact cagggcgcaa gggctgctaa aggaagcgga acacgtagaa
     1801 agccagtccg cagaaacggt gctgaccccg gatgaatgtc agctactggg ctatctggac
     1861 aagggaaaac gcaagcgcaa agagaaagca ggtagcttgc agtgggctta catggcgata
     1921 gctagactgg gcggttttat ggacagcaag cgaaccggaa ttgccagctg gggcgccctc
     1981 tgggaaggtt gggaagccct gcaaagtaaa ctggatggct ttcttgccgc caaggatctg
     2041 atggcgcagg ggatcaagat cctttaataa ggatccggct gctaacaaag cccgaaagga
     2101 agctgagttg gctgctgcca ccgctgagca ataactagca taaccccttg gggcctctaa
     2161 acgggtcttg aggggttttt tgctgaaagg aggaactata tccggatatc ccgcaagagg
     2221 cccggcagta ccggcataac caagcctatg cctacagcat ccagggtgac ggtgccgagg
     2281 atgacgatga gcgcattgtt agatttcata cacggtgcct gactgcgtta gcaatttaac
     2341 tgtgataaac taccgcatta aagcttatcg atgataagct gtcaaacatg agaattcttg
     2401 aagacgaaag ggcctcgtga tacgcctatt tttataggtt aatgtcatga taataatggt
     2461 ttcttagacg tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt
     2521 tttctaaata cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca
     2581 ataatattga aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt
     2641 ttttgcggca ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga
     2701 tgctgaagat cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa
     2761 gatccttgag agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct
     2821 gctatgtggc gcggtattat cccgtgttga cgccgggcaa gagcaactcg gtcgccgcat
     2881 acactattct cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga
     2941 tggcatgaca gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc
     3001 caacttactt ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat
     3061 gggggatcat gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa
     3121 cgacgagcgt gacaccacga tgcctgcagc aatggcaaca acgttgcgca aactattaac
     3181 tggcgaacta cttactctag cttcccggca acaattaata gactggatgg aggcggataa
     3241 agttgcagga ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc
     3301 tggagccggt gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc
     3361 ctcccgtatc gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag
     3421 acagatcgct gagataggtg cctcactgat taagcattgg taactgtcag accaagttta
     3481 ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa
     3541 gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc
     3601 gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat
     3661 ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga
     3721 gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt
     3781 ccttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata
     3841 cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac
     3901 cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg
     3961 ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg
     4021 tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag
     4081 cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct
     4141 ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc
     4201 aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt
     4261 ttgctggcct tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg
     4321 tattaccgcc tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga
     4381 gtcagtgagc gaggaagcgg aagagcgcct gatgcggtat tttctcctta cgcatctgtg
     4441 cggtatttca caccgcaatg gtgcactctc agtacaatct gctctgatgc cgcatagtta
     4501 agccagtata cactccgcta tcgctacgtg actgggtcat ggctgcgccc cgacacccgc
     4561 caacacccgc tgacgcgccc tgacgggctt gtctgctccc ggcatccgct tacagacaag
     4621 ctgtgaccgt ctccgggagc tgcatgtgtc agaggttttc accgtcatca ccgaaacgcg
     4681 cgaggcagct gcggtaaagc tcatcagcgt ggtcgtgaag cgattcacag atgtctgcct
     4741 gttcatccgc gtccagctcg ttgagtttct ccagaagcgt taatgtctgg cttctgataa
     4801 agcgggccat gttaagggcg gttttttcct gtttggtcac tgatgcctcc gtgtaagggg
     4861 gatttctgtt catgggggta atgataccga tgaaacgaga gaggatgctc acgatacggg
     4921 ttactgatga tgaacatgcc cggttactgg aacgttgtga gggtaaacaa ctggcggtat
     4981 ggatgcggcg ggaccagaga aaaatcactc agggtcaatg ccagcgcttc gttaatacag
     5041 atgtaggtgt tccacagggt agccagcagc atcctgcgat gcagatccgg aacataatgg
     5101 tgcagggcgc tgacttccgc gtttccagac tttacgaaac acggaaaccg aagaccattc
     5161 atgttgttgc tcaggtcgca gacgttttgc agcagcagtc gcttcacgtt cgctcgcgta
     5221 tcggtgattc attctgctaa ccagtaaggc aaccccgcca gcctagccgg gtcctcaacg
     5281 acaggagcac gatcatgcgc acccgtggcc aggacccaac gctgcccgag atgcgccgcg
     5341 tgcggctgct ggagatggcg gacgcgatgg atatgttctg ccaagggttg gtttgcgcat
     5401 tcacagttct ccgcaagaat tgattggctc caattcttgg agtggtgaat ccgttagcga
     5461 ggtgccgccg gcttccattc aggtcgaggt ggcccggctc catgcaccgc gacgcaacgc
     5521 ggggaggcag acaaggtata gggcggcgcc tacaatccat gccaacccgt tccatgtgct
     5581 cgccgaggcg gcataaatcg ccgtgacgat cagcggtcca atgatcgaag ttaggctggt
     5641 aagagccgcg agcgatcctt gaagctgtcc ctgatggtcg tcatctacct gcctggacag
     5701 catggcctgc aacgcgggca tcccgatgcc gccggaagcg agaagaatca taatggggaa
     5761 ggccatccag cctcgcgtcg cgaacgccag caagacgtag cccagcgcgt cggccgccat
     5821 gccggcgata atggcctgct tctcgccgaa acgtttggtg gcgggaccag tgacgaaggc
     5881 ttgagcgagg gcgtgcaaga ttccgaatac cgcaagcgac aggccgatca tcgtcgcgct
     5941 ccagcgaaag cggtcctcgc cgaaaatgac ccagagcgct gccggcacct gtcctacgag
     6001 ttgcatgata aagaagacag tcataagtgc ggcgacgata gtcatgcccc gcgcccaccg
     6061 gaaggagctg actgggttga aggctctcaa gggcatcggt cgagatcccg gtgcctaatg
     6121 agtgagctaa cttacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct
     6181 gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg
     6241 gcgccagggt ggtttttctt ttcaccagtg agacgggcaa cagctgattg cccttcaccg
     6301 cctggccctg agagagttgc agcaagcggt ccacgctggt ttgccccagc aggcgaaaat
     6361 cctgtttgat ggtggttaac ggcgggatat aacatgagct gtcttcggta tcgtcgtatc
     6421 ccactaccga gatatccgca ccaacgcgca gcccggactc ggtaatggcg cgcattgcgc
     6481 ccagcgccat ctgatcgttg gcaaccagca tcgcagtggg aacgatgccc tcattcagca
     6541 tttgcatggt ttgttgaaaa ccggacatgg cactccagtc gccttcccgt tccgctatcg
     6601 gctgaatttg attgcgagtg agatatttat gccagccagc cagacgcaga cgcgccgaga
     6661 cagaacttaa tgggcccgct aacagcgcga tttgctggtg acccaatgcg accagatgct
     6721 ccacgcccag tcgcgtaccg tcttcatggg agaaaataat actgttgatg ggtgtctggt
     6781 cagagacatc aagaaataac gccggaacat tagtgcaggc agcttccaca gcaatggcat
     6841 cctggtcatc cagcggatag ttaatgatca gcccactgac gcgttgcgcg agaagattgt
     6901 gcaccgccgc tttacaggct tcgacgccgc ttcgttctac catcgacacc accacgctgg
     6961 cacccagttg atcggcgcga gatttaatcg ccgcgacaat ttgcgacggc gcgtgcaggg
     7021 ccagactgga ggtggcaacg ccaatcagca acgactgttt gcccgccagt tgttgtgcca
     7081 cgcggttggg aatgtaattc agctccgcca tcgccgcttc cactttttcc cgcgttttcg
     7141 cagaaacgtg gctggcctgg ttcaccacgc gggaaacggt ctgataagag acaccggcat
     7201 actctgcgac atcgtataac gttactggtt tcacattcac caccctgaat tgactctctt
     7261 ccgggcgcta tcatgccata ccgcgaaagg ttttgcgcca ttcgatggtg tccgggatct
     7321 cgacgctctc ccttatgcga ctcctgcatt aggaagcagc ccagtagtag gttgaggccg
     7381 ttgagcaccg ccgccgcaag gaatggtgca tgcaaggaga tggcgcccaa cagtcccccg
     7441 gccacggggc ctgccaccat acccacgccg aaacaagcgc tcatgagccc gaagtggcga
     7501 gcccgatctt ccccatcggt gatgtcggcg atataggcgc cagcaaccgc acctgtggcg
     7561 ccggtgatgc cggccacgat gcgtccggcg tagaggatcg agatctcgat cccgcgaaat
     7621 taatacgact cactataggg gaattgtgag cggataacaa ttcccctcta gaaataattt
     7681 tgtttaactt taagaaggag a
//
