LOCUS       sequence-455395-        6321 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  10xHis-SUMO (Smt3) tagged HyperTEV60 engineered protease (E. coli 
            based protein expression).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6321)
  TITLE     Molecular Toolbox
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6321)
  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            "Unpublished"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: pHyperTEV60
FEATURES             Location/Qualifiers
     source          1..6321
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     193..212
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     terminator      complement(336..383)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     379..397
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(450..467)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(983..994)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     CDS             complement(1529..1558)
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     RBS             1572..1594
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    1609..1633
                     /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     complement(1633..1652)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1634..1652)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1736..1755)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        1961..2038
                     /gene="lacI"
                     /label=lacI promoter
     CDS             2039..3121
                     /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(2058..2077)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             3930..4121
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(4115..4137)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4223..4362
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4278..4297
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4377..4394)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4548..5136)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4628..4647)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(5247..5266)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             5258..6073
                     /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             5258..6073
                     /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     complement(5331..5350)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      complement(join(6165..6321,1..299))
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(join(6302..6321,1..2))
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 actattaaag aacgtggact ccaacgtcaa agggcgaaaa accgtctatc agggcgatgg
       61 cccactacgt gaaccatcac cctaatcaag ttttttgggg tcgaggtgcc gtaaagcact
      121 aaatcggaac cctaaaggga gcccccgatt tagagcttga cggggaaagc cggcgaacgt
      181 ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct agggcgctgg caagtgtagc
      241 ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat gcgccgctac agggcgcgtc
      301 ccattcgcca atccggatat agttcctcct ttcagcaaaa aacccctcaa gacccgttta
      361 gaggccccaa ggggttatgc tagttattgc tcagcggtgg cagcagccaa ctcagcttcc
      421 tttcgggctt tgttagcagc cggatctcag tggtggtggt ggtggtgctc gactaagttg
      481 gcagcatcac ccgacgcact ttgcgccgaa taaataccta agcttgtctt cactcgaggg
      541 tacccatgga tccttattac ggtttatcca taaacacttt atggccgccc cattccacgc
      601 tatcgctgtt caggctccag ccgctcaccc atttctgcag gctcggatcg gtcagcaggc
      661 gcataaaatc cggcggcacc gcggtaaaat agttgttggt gttggtaaag ttgctcgcgc
      721 tatgaatgcc cacaatgctg ccatcggtca cgctcaccat cgggctgccg cactggccat
      781 ctttcgtcgg aatccaatgt ttccaaaaca cgccatcgga gctcggatag gtggtgctca
      841 catcgctcac ttcgctggtc ggggttttgg tctgaaagtt gcgggtcacc agcacaattt
      901 cttcgcccac caccggttca cgaaacacca ggttggtcgg aaacggcgga aaatctttcg
      961 gcatttcaat cagcaccaga tcgcggcctt cgatcagatg cagtttcagg gtggtggtgt
     1021 tgctaatggt aaaggtgcca tgtttgctgg taatggtcag ggtgccgttg ttgcggcgaa
     1081 acaggtgcgc gttggtaata atcagcgggc caaagccaat gccatacagg ctaatgctat
     1141 agccatcgct ggtattggtc agcagaacaa tggtatcgct aatcgggtta taatcgcgcg
     1201 ggcccggcgc cgcgctttcc gcggatccgc cgccaatctg ttcacgatgc gcttcaataa
     1261 tatcgttatc ttccatatcc agatcttccg gggtctgatc cgcctgaata cgaatgccat
     1321 catacagaaa acgcaggcta tccatttctt tgccctgacg tttcgcaaac gcttccatca
     1381 gacgacgcag cggggtggtt tttttaattt taaaaaaaat ttcgctgctg ccatcgctca
     1441 ctttcaggtt aatatgggtt tccggtttca cttccggttt cacttccggt ttcgcttcct
     1501 ggttcacttc gctatcgctc atgctgccgt gatggtgatg gtgatggtga tggtgatggc
     1561 ccatatgtat atctccttct taaagttaaa caaaattatt tctagagggg aattgttatc
     1621 cgctcacaat tcccctatag tgagtcgtat taatttcgcg ggatcgagat ctcgatcctc
     1681 tacgccggac gcatcgtggc cggcatcacc ggcgccacag gtgcggttgc tggcgcctat
     1741 atcgccgaca tcaccgatgg ggaagatcgg gctcgccact tcgggctcat gagcgcttgt
     1801 ttcggcgtgg gtatggtggc aggccccgtg gccgggggac tgttgggcgc catctccttg
     1861 catgcaccat tccttgcggc ggcggtgctc aacggcctca acctactact gggctgcttc
     1921 ctaatgcagg agtcgcataa gggagagcgt cgagatcccg gacaccatcg aatggcgcaa
     1981 aacctttcgc ggtatggcat gatagcgccc ggaagagagt caattcaggg tggtgaatgt
     2041 gaaaccagta acgttatacg atgtcgcaga gtatgccggt gtctcttatc agaccgtttc
     2101 ccgcgtggtg aaccaggcca gccacgtttc tgcgaaaacg cgggaaaaag tggaagcggc
     2161 gatggcggag ctgaattaca ttcccaaccg cgtggcacaa caactggcgg gcaaacagtc
     2221 gttgctgatt ggcgttgcca cctccagtct ggccctgcac gcgccgtcgc aaattgtcgc
     2281 ggcgattaaa tctcgcgccg atcaactggg tgccagcgtg gtggtgtcga tggtagaacg
     2341 aagcggcgtc gaagcctgta aagcggcggt gcacaatctt ctcgcgcaac gcgtcagtgg
     2401 gctgatcatt aactatccgc tggatgacca ggatgccatt gctgtggaag ctgcctgcac
     2461 taatgttccg gcgttatttc ttgatgtctc tgaccagaca cccatcaaca gtattatttt
     2521 ctcccatgaa gacggtacgc gactgggcgt ggagcatctg gtcgcattgg gtcaccagca
     2581 aatcgcgctg ttagcgggcc cattaagttc tgtctcggcg cgtctgcgtc tggctggctg
     2641 gcataaatat ctcactcgca atcaaattca gccgatagcg gaacgggaag gcgactggag
     2701 tgccatgtcc ggttttcaac aaaccatgca aatgctgaat gagggcatcg ttcccactgc
     2761 gatgctggtt gccaacgatc agatggcgct gggcgcaatg cgcgccatta ccgagtccgg
     2821 gctgcgcgtt ggtgcggata tctcggtagt gggatacgac gataccgaag acagctcatg
     2881 ttatatcccg ccgttaacca ccatcaaaca ggattttcgc ctgctggggc aaaccagcgt
     2941 ggaccgcttg ctgcaactct ctcagggcca ggcggtgaag ggcaatcagc tgttgcccgt
     3001 ctcactggtg aaaagaaaaa ccaccctggc gcccaatacg caaaccgcct ctccccgcgc
     3061 gttggccgat tcattaatgc agctggcacg acaggtttcc cgactggaaa gcgggcagtg
     3121 agcgcaacgc aattaatgta agttagctca ctcattaggc accgggatct cgaccgatgc
     3181 ccttgagagc cttcaaccca gtcagctcct tccggtgggc gcggggcatg actatcgtcg
     3241 ccgcacttat gactgtcttc tttatcatgc aactcgtagg acaggtgccg gcagcgctct
     3301 gggtcatttt cggcgaggac cgctttcgct ggagcgcgac gatgatcggc ctgtcgcttg
     3361 cggtattcgg aatcttgcac gccctcgctc aagccttcgt cactggtccc gccaccaaac
     3421 gtttcggcga gaagcaggcc attatcgccg gcatggcggc cccacgggtg cgcatgatcg
     3481 tgctcctgtc gttgaggacc cggctaggct ggcggggttg ccttactggt tagcagaatg
     3541 aatcaccgat acgcgagcga acgtgaagcg actgctgctg caaaacgtct gcgacctgag
     3601 caacaacatg aatggtcttc ggtttccgtg tttcgtaaag tctggaaacg cggaagtcag
     3661 cgccctgcac cattatgttc cggatctgca tcgcaggatg ctgctggcta ccctgtggaa
     3721 cacctacatc tgtattaacg aagcgctggc attgaccctg agtgattttt ctctggtccc
     3781 gccgcatcca taccgccagt tgtttaccct cacaacgttc cagtaaccgg gcatgttcat
     3841 catcagtaac ccgtatcgtg agcatcctct ctcgtttcat cggtatcatt acccccatga
     3901 acagaaatcc cccttacacg gaggcatcag tgaccaaaca ggaaaaaacc gcccttaaca
     3961 tggcccgctt tatcagaagc cagacattaa cgcttctgga gaaactcaac gagctggacg
     4021 cggatgaaca ggcagacatc tgtgaatcgc ttcacgacca cgctgatgag ctttaccgca
     4081 gctgcctcgc gcgtttcggt gatgacggtg aaaacctctg acacatgcag ctcccggaga
     4141 cggtcacagc ttgtctgtaa gcggatgccg ggagcagaca agcccgtcag ggcgcgtcag
     4201 cgggtgttgg cgggtgtcgg ggcgcagcca tgacccagtc acgtagcgat agcggagtgt
     4261 atactggctt aactatgcgg catcagagca gattgtactg agagtgcacc attgcggtgt
     4321 gaaataccgc acagatgcgt aaggagaaaa taccgcatca ggcgctcttc cgcttcctcg
     4381 ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag
     4441 gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa
     4501 ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc
     4561 cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca
     4621 ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg
     4681 accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct
     4741 catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt
     4801 gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag
     4861 tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc
     4921 agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac
     4981 actagaagga cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga
     5041 gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc
     5101 aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg
     5161 gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gaacaataaa
     5221 actgtctgct tacataaaca gtaatacaag gggtgttatg agccatattc aacgggaaac
     5281 gtcttgctct aggccgcgat taaattccaa catggatgct gatttatatg ggtataaatg
     5341 ggctcgcgat aatgtcgggc aatcaggtgc gacaatctat cgattgtatg ggaagcccga
     5401 tgcgccagag ttgtttctga aacatggcaa aggtagcgtt gccaatgatg ttacagatga
     5461 gatggtcaga ctaaactggc tgacggaatt tatgcctctt ccgaccatca agcattttat
     5521 ccgtactcct gatgatgcat ggttactcac cactgcgatc cccgggaaaa cagcattcca
     5581 ggtattagaa gaatatcctg attcaggtga aaatattgtt gatgcgctgg cagtgttcct
     5641 gcgccggttg cattcgattc ctgtttgtaa ttgtcctttt aacagcgatc gcgtatttcg
     5701 tctcgctcag gcgcaatcac gaatgaataa cggtttggtt gatgcgagtg attttgatga
     5761 cgagcgtaat ggctggcctg ttgaacaagt ctggaaagaa atgcataaac ttttgccatt
     5821 ctcaccggat tcagtcgtca ctcatggtga tttctcactt gataacctta tttttgacga
     5881 ggggaaatta ataggttgta ttgatgttgg acgagtcgga atcgcagacc gataccagga
     5941 tcttgccatc ctatggaact gcctcggtga gttttctcct tcattacaga aacggctttt
     6001 tcaaaaatat ggtattgata atcctgatat gaataaattg cagtttcatt tgatgctcga
     6061 tgagtttttc taagaattaa ttcatgagcg gatacatatt tgaatgtatt tagaaaaata
     6121 aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctaaattg taagcgttaa
     6181 tattttgtta aaattcgcgt taaatttttg ttaaatcagc tcatttttta accaataggc
     6241 cgaaatcggc aaaatccctt ataaatcaaa agaatagacc gagatagggt tgagtgttgt
     6301 tccagtttgg aacaagagtc c
//
