LOCUS       sequence-386265-        6036 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  protein expression of the His-tagged catalytic domain.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6036)
  TITLE     Knapp lab plasmids
REFERENCE   2  (bases 1 to 6036)
  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"
COMMENT     Sequence Label: pNIC28-SENP1
FEATURES             Location/Qualifiers
     source          1..6036
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     45..67
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(61..252)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(1061..2143)
                     /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     2105..2124
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(2144..2221)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     2427..2446
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     2530..2549
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2530..2548
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    2549..2573
                     /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             2588..2610
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             2621..2638
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             2663..2683
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             3430..3447
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(3500..3518)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      3514..3561
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      3598..4053
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3685..3704)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3895..3916
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(4145..4960)
                     /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(4145..4960)
                     /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     4868..4887
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     4952..4971
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      5082..5670
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5571..5590
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5824..5841
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    5856..5995
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(5921..5940)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 ttgtctgctc ccggcatccg cttacagaca agctgtgacc gtctccggga gctgcatgtg
       61 tcagaggttt tcaccgtcat caccgaaacg cgcgaggcag ctgcggtaaa gctcatcagc
      121 gtggtcgtga agcgattcac agatgtctgc ctgttcatcc gcgtccagct cgttgagttt
      181 ctccagaagc gttaatgtct ggcttctgat aaagcgggcc atgttaaggg cggttttttc
      241 ctgtttggtc actgatgcct ccgtgtaagg gggatttctg ttcatggggg taatgatacc
      301 gatgaaacga gagaggatgc tcacgatacg ggttactgat gatgaacatg cccggttact
      361 ggaacgttgt gagggtaaac aactggcggt atggatgcgg cgggaccaga gaaaaatcac
      421 tcagggtcaa tgccagcgct tcgttaatac agatgtaggt gttccacagg gtagccagca
      481 gcatcctgcg atgcagatcc ggaacataat ggtgcagggc gctgacttcc gcgtttccag
      541 actttacgaa acacggaaac cgaagaccat tcatgttgtt gctcaggtcg cagacgtttt
      601 gcagcagcag tcgcttcacg ttcgctcgcg tatcggtgat tcattctgct aaccagtaag
      661 gcaaccccgc cagcctagcc gggtcctcaa cgacaggagc acgatcatgc gcacccgtgg
      721 ggccgccatg ccggcgataa tggcctgctt ctcgccgaaa cgtttggtgg cgggaccagt
      781 gacgaaggct tgagcgaggg cgtgcaagat tccgaatacc gcaagcgaca ggccgatcat
      841 cgtcgcgctc cagcgaaagc ggtcctcgcc gaaaatgacc cagagcgctg ccggcacctg
      901 tcctacgagt tgcatgataa agaagacagt cataagtgcg gcgacgatag tcatgccccg
      961 cgcccaccgg aaggagctga ctgggttgaa ggctctcaag ggcatcggtc gagatcccgg
     1021 tgcctaatga gtgagctaac ttacattaat tgcgttgcgc tcactgcccg ctttccagtc
     1081 gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt
     1141 gcgtattggg cgccagggtg gtttttcttt tcaccagtga gacgggcaac agctgattgc
     1201 ccttcaccgc ctggccctga gagagttgca gcaagcggtc cacgctggtt tgccccagca
     1261 ggcgaaaatc ctgtttgatg gtggttaacg gcgggatata acatgagctg tcttcggtat
     1321 cgtcgtatcc cactaccgag atatccgcac caacgcgcag cccggactcg gtaatggcgc
     1381 gcattgcgcc cagcgccatc tgatcgttgg caaccagcat cgcagtggga acgatgccct
     1441 cattcagcat ttgcatggtt tgttgaaaac cggacatggc actccagtcg ccttcccgtt
     1501 ccgctatcgg ctgaatttga ttgcgagtga gatatttatg ccagccagcc agacgcagac
     1561 gcgccgagac agaacttaat gggcccgcta acagcgcgat ttgctggtga cccaatgcga
     1621 ccagatgctc cacgcccagt cgcgtaccgt cttcatggga gaaaataata ctgttgatgg
     1681 gtgtctggtc agagacatca agaaataacg ccggaacatt agtgcaggca gcttccacag
     1741 caatggcatc ctggtcatcc agcggatagt taatgatcag cccactgacg cgttgcgcga
     1801 gaagattgtg caccgccgct ttacaggctt cgacgccgct tcgttctacc atcgacacca
     1861 ccacgctggc acccagttga tcggcgcgag atttaatcgc cgcgacaatt tgcgacggcg
     1921 cgtgcagggc cagactggag gtggcaacgc caatcagcaa cgactgtttg cccgccagtt
     1981 gttgtgccac gcggttggga atgtaattca gctccgccat cgccgcttcc actttttccc
     2041 gcgttttcgc agaaacgtgg ctggcctggt tcaccacgcg ggaaacggtc tgataagaga
     2101 caccggcata ctctgcgaca tcgtataacg ttactggttt cacattcacc accctgaatt
     2161 gactctcttc cgggcgctat catgccatac cgcgaaaggt tttgcgccat tcgatggtgt
     2221 ccgggatctc gacgctctcc cttatgcgac tcctgcatta ggaagcagcc cagtagtagg
     2281 ttgaggccgt tgagcaccgc cgccgcaagg aatggtgcat gcaaggagat ggcgcccaac
     2341 agtcccccgg ccacggggcc tgccaccata cccacgccga aacaagcgct catgagcccg
     2401 aagtggcgag cccgatcttc cccatcggtg atgtcggcga tataggcgcc agcaaccgca
     2461 cctgtggcgc cggtgatgcc ggccacgatg cgtccggcgt agaggatcga gatctcgatc
     2521 ccgcgaaatt aatacgactc actatagggg aattgtgagc ggataacaat tcccctctag
     2581 aaataatttt gtttaacttt aagaaggaga tatacatatg caccatcatc atcatcattc
     2641 ttctggtgta gatctgggta ccgagaacct gtacttccaa tccatggaat ttcctgaaat
     2701 tacagaggaa atggagaaag aaataaagaa tgtatttcgt aatgggaatc aggatgaagt
     2761 tctcagtgaa gcatttcgcc tgaccattac acgcaaagat attcaaactc taaaccatct
     2821 gaattggctc aatgatgaga tcatcaattt ctacatgaat atgctgatgg agcgaagtaa
     2881 agagaagggc ttgccaagtg tgcatgcatt taataccttt ttcttcacta aattaaaaac
     2941 ggctggttat caggcagtga aacgttggac aaagaaagta gatgtatttt ctgttgacat
     3001 tcttttggtg cccattcacc tgggagtaca ctggtgtcta gctgttgtgg actttagaaa
     3061 gaagaatatt acctattacg actccatggg tgggataaac aatgaagcct gcagaatact
     3121 cttgcaatac ctaaagcaag aaagcattga caagaaaagg aaagagtttg acaccaatgg
     3181 ctggcagctt ttcagcaaga aaagccagga gattcctcag cagatgaatg gaagtgactg
     3241 tgggatgttt gcctgcaaat atgctgactg tattaccaaa gacagaccaa tcaacttcac
     3301 acagcaacac atgccatact tccggaagcg gatggtctgg gagatcctcc accgaaaact
     3361 cttgtaacag taaaggtgga tacggatccg aattcgagct ccgtcgacaa gcttgcggcc
     3421 gcactcgagc accaccacca ccaccactga gatccggctg ctaacaaagc ccgaaaggaa
     3481 gctgagttgg ctgctgccac cgctgagcaa taactagcat aaccccttgg ggcctctaaa
     3541 cgggtcttga ggggtttttt gctgaaagga ggaactatat ccggattggc gaatgggacg
     3601 cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta
     3661 cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt
     3721 tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg
     3781 ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat
     3841 cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac
     3901 tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag
     3961 ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg
     4021 cgaattttaa caaaatatta acgcttacaa tttaggtggc acttttcggg gaaatgtgcg
     4081 cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgaatta
     4141 attcttagaa aaactcatcg agcatcaaat gaaactgcaa tttattcata tcaggattat
     4201 caataccata tttttgaaaa agccgtttct gtaatgaagg agaaaactca ccgaggcagt
     4261 tccataggat ggcaagatcc tggtatcggt ctgcgattcc gactcgtcca acatcaatac
     4321 aacctattaa tttcccctcg tcaaaaataa ggttatcaag tgagaaatca ccatgagtga
     4381 cgactgaatc cggtgagaat ggcaaaagtt tatgcatttc tttccagact tgttcaacag
     4441 gccagccatt acgctcgtca tcaaaatcac tcgcatcaac caaaccgtta ttcattcgtg
     4501 attgcgcctg agcgagacga aatacgcgat cgctgttaaa aggacaatta caaacaggaa
     4561 tcgaatgcaa ccggcgcagg aacactgcca gcgcatcaac aatattttca cctgaatcag
     4621 gatattcttc taatacctgg aatgctgttt tcccggggat cgcagtggtg agtaaccatg
     4681 catcatcagg agtacggata aaatgcttga tggtcggaag aggcataaat tccgtcagcc
     4741 agtttagtct gaccatctca tctgtaacat cattggcaac gctacctttg ccatgtttca
     4801 gaaacaactc tggcgcatcg ggcttcccat acaatcgata gattgtcgca cctgattgcc
     4861 cgacattatc gcgagcccat ttatacccat ataaatcagc atccatgttg gaatttaatc
     4921 gcggcctaga gcaagacgtt tcccgttgaa tatggctcat aacacccctt gtattactgt
     4981 ttatgtaagc agacagtttt attgttcatg accaaaatcc cttaacgtga gttttcgttc
     5041 cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg
     5101 cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg
     5161 gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca
     5221 aatactgtcc ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg
     5281 cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg
     5341 tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga
     5401 acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac
     5461 ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat
     5521 ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc
     5581 tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga
     5641 tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc
     5701 ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg
     5761 gataaccgta ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag
     5821 cgcagcgagt cagtgagcga ggaagcggaa gagcgcctga tgcggtattt tctccttacg
     5881 catctgtgcg gtatttcaca ccgcaatggt gcactctcag tacaatctgc tctgatgccg
     5941 catagttaag ccagtataca ctccgctatc gctacgtgac tgggtcatgg ctgcgccccg
     6001 acacccgcca acacccgctg acgcgccctg acgggc
//
