LOCUS       sequence-376453-        6246 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses P. horikoshii tetrahedral aminopeptidase 2 (TET2) in E. 
            coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6246)
  AUTHORS   Gauto DF, Macek P, Malinverni D, Fraga H, Paloni M, Sucec I, Hessel 
            A, Bustamante JP, Barducci A, Schanda P
  TITLE     Functional control of a 0.5 MDa TET aminopeptidase by a flexible 
            loop revealed by MAS NMR.
  JOURNAL   Nat Commun. 2022 Apr 8;13(1):1927. doi: 10.1038/s41467-022-29423-0.
  PUBMED    35395851
REFERENCE   2  (bases 1 to 6246)
  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: "Nat 
            Commun."; date: "2022-04-8"; volume: "13(1):1927. doi"; pages: " 
            10.1038/s41467-022-29423-0"
COMMENT     Sequence Label: pET41c-PhTET2
FEATURES             Location/Qualifiers
     source          1..6246
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             69..92
                     /codon_start=1
                     /product="8xHis affinity tag"
                     /label=8xHis
                     /translation="HHHHHHHH"
     primer_bind     complement(155..173)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      169..216
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      253..708
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(340..359)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     550..571
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(800..1615)
                     /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(800..1615)
                     /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     1523..1542
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     1607..1626
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      1737..2325
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2226..2245
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2479..2496
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    2511..2650
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(2576..2595)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2736..2758
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(2752..2943)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(3515..4597)
                     /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     4559..4578
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(4598..4675)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     4881..4900
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     4988..5007
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4988..5006
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    5007..5031
                     /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             5046..5068
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
ORIGIN
        1 cgaattctgt acaggccttg gcgcgcctgc aggcgagctc cgtcgacaag cttgcggccg
       61 cactcgagca ccaccaccac caccaccacc actaattgat taatacctag gctgctaaac
      121 aaagcccgaa aggaagctga gttggctgct gccaccgctg agcaataact agcataaccc
      181 cttggggcct ctaaacgggt cttgaggggt tttttgctga aaggaggaac tatatccgga
      241 ttggcgaatg ggacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc
      301 gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt
      361 cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag
      421 ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag ggtgatggtt
      481 cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt
      541 tctttaatag tggactcttg ttccaaactg gaacaacact caaccctatc tcggtctatt
      601 cttttgattt ataagggatt ttgccgattt cggcctattg gttaaaaaat gagctgattt
      661 aacaaaaatt taacgcgaat tttaacaaaa tattaacgct tacaatttag gtggcacttt
      721 tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta
      781 tccgctcatg aattaattct tagaaaaact catcgagcat caaatgaaac tgcaatttat
      841 tcatatcagg attatcaata ccatattttt gaaaaagccg tttctgtaat gaaggagaaa
      901 actcaccgag gcagttccat aggatggcaa gatcctggta tcggtctgcg attccgactc
      961 gtccaacatc aatacaacct attaatttcc cctcgtcaaa aataaggtta tcaagtgaga
     1021 aatcaccatg agtgacgact gaatccggtg agaatggcaa aagtttatgc atttctttcc
     1081 agacttgttc aacaggccag ccattacgct cgtcatcaaa atcactcgca tcaaccaaac
     1141 cgttattcat tcgtgattgc gcctgagcga gacgaaatac gcgatcgctg ttaaaaggac
     1201 aattacaaac aggaatcgaa tgcaaccggc gcaggaacac tgccagcgca tcaacaatat
     1261 tttcacctga atcaggatat tcttctaata cctggaatgc tgttttcccg gggatcgcag
     1321 tggtgagtaa ccatgcatca tcaggagtac ggataaaatg cttgatggtc ggaagaggca
     1381 taaattccgt cagccagttt agtctgacca tctcatctgt aacatcattg gcaacgctac
     1441 ctttgccatg tttcagaaac aactctggcg catcgggctt cccatacaat cgatagattg
     1501 tcgcacctga ttgcccgaca ttatcgcgag cccatttata cccatataaa tcagcatcca
     1561 tgttggaatt taatcgcggc ctagagcaag acgtttcccg ttgaatatgg ctcataacac
     1621 cccttgtatt actgtttatg taagcagaca gttttattgt tcatgaccaa aatcccttaa
     1681 cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga
     1741 gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg
     1801 gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc
     1861 agagcgcaga taccaaatac tgtccttcta gtgtagccgt agttaggcca ccacttcaag
     1921 aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc
     1981 agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg
     2041 cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac
     2101 accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga
     2161 aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt
     2221 ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag
     2281 cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg
     2341 gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta
     2401 tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc
     2461 agccgaacga ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg cctgatgcgg
     2521 tattttctcc ttacgcatct gtgcggtatt tcacaccgca atggtgcact ctcagtacaa
     2581 tctgctctga tgccgcatag ttaagccagt atacactccg ctatcgctac gtgactgggt
     2641 catggctgcg ccccgacacc cgccaacacc cgctgacgcg ccctgacggg cttgtctgct
     2701 cccggcatcc gcttacagac aagctgtgac cgtctccggg agctgcatgt gtcagaggtt
     2761 ttcaccgtca tcaccgaaac gcgcgaggca gctgcggtaa agctcatcag cgtggtcgtg
     2821 aagcgattca cagatgtctg cctgttcatc cgcgtccagc tcgttgagtt tctccagaag
     2881 cgttaatgtc tggcttctga taaagcgggc catgttaagg gcggtttttt cctgtttggt
     2941 cactgatgcc tccgtgtaag ggggatttct gttcatgggg gtaatgatac cgatgaaacg
     3001 agagaggatg ctcacgatac gggttactga tgatgaacat gcccggttac tggaacgttg
     3061 tgagggtaaa caactggcgg tatggatgcg gcgggaccag agaaaaatca ctcagggtca
     3121 atgccagcgc ttcgttaata cagatgtagg tgttccacag ggtagccagc agcatcctgc
     3181 gatgcagatc cggaacataa tggtgcaggg cgctgacttc cgcgtttcca gactttacga
     3241 aacacggaaa ccgaagacca ttcatgttgt tgctcaggtc gcagacgttt tgcagcagca
     3301 gtcgcttcac gttcgctcgc gtatcggtga ttcattctgc taaccagtaa ggcaaccccg
     3361 ccagcctagc cgggtcctca acgacaggag cacgatcatg ctagtcatgc cccgcgccca
     3421 ccggaaggag ctgactgggt tgaaggctct caagggcatc ggtcgagatc ccggtgccta
     3481 atgagtgagc taacttacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa
     3541 cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat
     3601 tgggcgccag ggtggttttt cttttcacca gtgagacggg caacagctga ttgcccttca
     3661 ccgcctggcc ctgagagagt tgcagcaagc ggtccacgct ggtttgcccc agcaggcgaa
     3721 aatcctgttt gatggtggtt aacggcggga tataacatga gctgtcttcg gtatcgtcgt
     3781 atcccactac cgagatgtcc gcaccaacgc gcagcccgga ctcggtaatg gcgcgcattg
     3841 cgcccagcgc catctgatcg ttggcaacca gcatcgcagt gggaacgatg ccctcattca
     3901 gcatttgcat ggtttgttga aaaccggaca tggcactcca gtcgccttcc cgttccgcta
     3961 tcggctgaat ttgattgcga gtgagatatt tatgccagcc agccagacgc agacgcgccg
     4021 agacagaact taatgggccc gctaacagcg cgatttgctg gtgacccaat gcgaccagat
     4081 gctccacgcc cagtcgcgta ccgtcttcat gggagaaaat aatactgttg atgggtgtct
     4141 ggtcagagac atcaagaaat aacgccggaa cattagtgca ggcagcttcc acagcaatgg
     4201 catcctggtc atccagcgga tagttaatga tcagcccact gacgcgttgc gcgagaagat
     4261 tgtgcaccgc cgctttacag gcttcgacgc cgcttcgttc taccatcgac accaccacgc
     4321 tggcacccag ttgatcggcg cgagatttaa tcgccgcgac aatttgcgac ggcgcgtgca
     4381 gggccagact ggaggtggca acgccaatca gcaacgactg tttgcccgcc agttgttgtg
     4441 ccacgcggtt gggaatgtaa ttcagctccg ccatcgccgc ttccactttt tcccgcgttt
     4501 tcgcagaaac gtggctggcc tggttcacca cgcgggaaac ggtctgataa gagacaccgg
     4561 catactctgc gacatcgtat aacgttactg gtttcacatt caccaccctg aattgactct
     4621 cttccgggcg ctatcatgcc ataccgcgaa aggttttgcg ccattcgatg gtgtccggga
     4681 tctcgacgct ctcccttatg cgactcctgc attaggaagc agcccagtag taggttgagg
     4741 ccgttgagca ccgccgccgc aaggaatggt gcatgcaagg agatggcgcc caacagtccc
     4801 ccggccacgg ggcctgccac catacccacg ccgaaacaag cgctcatgag cccgaagtgg
     4861 cgagcccgat cttccccatc ggtgatgtcg gcgatatagg cgccagcaac cgcacctgtg
     4921 gcgccggtga tgccggccac gatgcgtccg gcgtagagga tcgagatcga tctcgatccc
     4981 gcgaaattaa tacgactcac tataggggaa ttgtgagcgg ataacaattc ccctctagaa
     5041 ataattttgt ttaactttaa gaaggagata tacatatgga ggtgaggaat atggtggact
     5101 acgagctttt aaaaaaggta gttgaagccc ctggagtttc tgggtatgaa ttcttgggga
     5161 ttagggatgt tgttatagag gagatcaagg actatgtaga tgaagttaag gttgacaagc
     5221 tgggaaacgt tatagctcac aagaagggag aagggccaaa agttatgata gcggcccaca
     5281 tggatcagat agggttgatg gttactcaca tagagaagaa cggattctta agagttgctc
     5341 cgattggagg agtcgatcca aaaactctga tagctcaaag gtttaaggtc tggattgata
     5401 aaggtaagtt catttatgga gttggcgcat cggtaccgcc tcacattcag aagccagagg
     5461 acaggaagaa ggcccctgat tgggatcaga tatttataga cataggtgct gaaagcaagg
     5521 aagaagctga ggatatggga gttaagatcg ggacggtaat tacgtgggat ggaagattgg
     5581 aaaggttagg taagcatagg ttcgtaagta tagcttttga tgatagaatt gcagtatata
     5641 cgatactgga agttgccaag cagcttaagg atgctaaggc tgatgtatac ttcgttgcca
     5701 ctgttcagga ggaggtagga cttagaggtg ctagaacgtc agcatttgga atcgaacccg
     5761 attatggctt tgcgattgat gttactatag cagcagacat tccaggaacg ccagagcaca
     5821 agcaagtaac ccacttggga aagggaactg cgataaagat aatggatcgc tcggttatat
     5881 gccatccaac gatagtaaga tggttggagg aattggcgaa gaagcatgag ataccatacc
     5941 agcttgagat cctcttaggt ggaggaacgg atgccggagc gatccactta acaaaagcag
     6001 gtgttccaac tggagcatta agcgttccag ctaggtatat tcactctaac actgaagtcg
     6061 ttgatgaaag agacgttgac gcaacggttg agctaatgac taaagccctt gaaaacatcc
     6121 atgagcttaa gatctaaacg tttatttacc tcttattttt accatatttc tggtgaattc
     6181 ataatgaggg ttattaagga atggaatgta aaaattaagc tagtaaggac caaaagaggc
     6241 gggatc
//
