LOCUS       sequence-414745-        5397 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression of the His-tagged peptide TREER..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5397)
  AUTHORS   Suyama A, Devlin KL, Macias-Contreras M, Doh JK, Shinde U, Beatty KE
  TITLE     Orthogonal Versatile Interacting Peptide Tags for Imaging Cellular 
            Proteins.
  JOURNAL   Biochemistry. 2023 Jun 6;62(11):1735-1743. doi: 
            10.1021/acs.biochem.2c00712. Epub 2023 May 11.
  PUBMED    37167569
REFERENCE   2  (bases 1 to 5397)
  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"; doi: 
            "10.1021/acs.biochem.2c00712"; journalName: "Biochemistry"; date: 
            "2023-06-6- 6"; volume: "62"; issue: "11"; pages: "1735-1743"
COMMENT     Sequence Label: pET28b-TREER
FEATURES             Location/Qualifiers
     source          1..5397
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     946..965
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(985..1062)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     1268..1287
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     1371..1390
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1371..1389
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1390..1414
                     /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             1429..1451
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             1632..1649
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1702..1720)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1716..1763
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1800..2255
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1887..1906)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2097..2118
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(2347..3162)
                     /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(2347..3162)
                     /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     3070..3089
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     3154..3173
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      3284..3872
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3773..3792
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4026..4043
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4058..4197
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4123..4142)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4283..4305
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4299..4490)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(join(5299..5397,1..984))
                     /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"
ORIGIN
        1 ggtttttctt ttcaccagtg agacgggcaa cagctgattg cccttcaccg cctggccctg
       61 agagagttgc agcaagcggt ccacgctggt ttgccccagc aggcgaaaat cctgtttgat
      121 ggtggttaac ggcgggatat aacatgagct gtcttcggta tcgtcgtatc ccactaccga
      181 gatatccgca ccaacgcgca gcccggactc ggtaatggcg cgcattgcgc ccagcgccat
      241 ctgatcgttg gcaaccagca tcgcagtggg aacgatgccc tcattcagca tttgcatggt
      301 ttgttgaaaa ccggacatgg cactccagtc gccttcccgt tccgctatcg gctgaatttg
      361 attgcgagtg agatatttat gccagccagc cagacgcaga cgcgccgaga cagaacttaa
      421 tgggcccgct aacagcgcga tttgctggtg acccaatgcg accagatgct ccacgcccag
      481 tcgcgtaccg tcttcatggg agaaaataat actgttgatg ggtgtctggt cagagacatc
      541 aagaaataac gccggaacat tagtgcaggc agcttccaca gcaatggcat cctggtcatc
      601 cagcggatag ttaatgatca gcccactgac gcgttgcgcg agaagattgt gcaccgccgc
      661 tttacaggct tcgacgccgc ttcgttctac catcgacacc accacgctgg cacccagttg
      721 atcggcgcga gatttaatcg ccgcgacaat ttgcgacggc gcgtgcaggg ccagactgga
      781 ggtggcaacg ccaatcagca acgactgttt gcccgccagt tgttgtgcca cgcggttggg
      841 aatgtaattc agctccgcca tcgccgcttc cactttttcc cgcgttttcg cagaaacgtg
      901 gctggcctgg ttcaccacgc gggaaacggt ctgataagag acaccggcat actctgcgac
      961 atcgtataac gttactggtt tcacattcac caccctgaat tgactctctt ccgggcgcta
     1021 tcatgccata ccgcgaaagg ttttgcgcca ttcgatggtg tccgggatct cgacgctctc
     1081 ccttatgcga ctcctgcatt aggaagcagc ccagtagtag gttgaggccg ttgagcaccg
     1141 ccgccgcaag gaatggtgca tgcaaggaga tggcgcccaa cagtcccccg gccacggggc
     1201 ctgccaccat acccacgccg aaacaagcgc tcatgagccc gaagtggcga gcccgatctt
     1261 ccccatcggt gatgtcggcg atataggcgc cagcaaccgc acctgtggcg ccggtgatgc
     1321 cggccacgat gcgtccggcg tagaggatcg agatctcgat cccgcgaaat taatacgact
     1381 cactataggg gaattgtgag cggataacaa ttcccctcta gaaataattt tgtttaactt
     1441 taagaaggag atataccatg ggttcttctc tggaaatccg tgttgcgttc ctgcgtcagg
     1501 aagtcaccgc gctggaaacc gaaaacgcgg aactggaaca gcgtgttcag cgtctgcgta
     1561 accgttacgg tccgctgggt ggtggtgcgg ctgcatgggg tctgtgctac ccgtgggttt
     1621 acggtctcga gcaccaccac caccaccact gagatccggc tgctaacaaa gcccgaaagg
     1681 aagctgagtt ggctgctgcc accgctgagc aataactagc ataacccctt ggggcctcta
     1741 aacgggtctt gaggggtttt ttgctgaaag gaggaactat atccggattg gcgaatggga
     1801 cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc
     1861 tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac
     1921 gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag
     1981 tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc
     2041 atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg
     2101 actcttgttc caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata
     2161 agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa
     2221 cgcgaatttt aacaaaatat taacgcttac aatttaggtg gcacttttcg gggaaatgtg
     2281 cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaat
     2341 taattcttag aaaaactcat cgagcatcaa atgaaactgc aatttattca tatcaggatt
     2401 atcaatacca tatttttgaa aaagccgttt ctgtaatgaa ggagaaaact caccgaggca
     2461 gttccatagg atggcaagat cctggtatcg gtctgcgatt ccgactcgtc caacatcaat
     2521 acaacctatt aatttcccct cgtcaaaaat aaggttatca agtgagaaat caccatgagt
     2581 gacgactgaa tccggtgaga atggcaaaag tttatgcatt tctttccaga cttgttcaac
     2641 aggccagcca ttacgctcgt catcaaaatc actcgcatca accaaaccgt tattcattcg
     2701 tgattgcgcc tgagcgagac gaaatacgcg atcgctgtta aaaggacaat tacaaacagg
     2761 aatcgaatgc aaccggcgca ggaacactgc cagcgcatca acaatatttt cacctgaatc
     2821 aggatattct tctaatacct ggaatgctgt tttcccgggg atcgcagtgg tgagtaacca
     2881 tgcatcatca ggagtacgga taaaatgctt gatggtcgga agaggcataa attccgtcag
     2941 ccagtttagt ctgaccatct catctgtaac atcattggca acgctacctt tgccatgttt
     3001 cagaaacaac tctggcgcat cgggcttccc atacaatcga tagattgtcg cacctgattg
     3061 cccgacatta tcgcgagccc atttataccc atataaatca gcatccatgt tggaatttaa
     3121 tcgcggccta gagcaagacg tttcccgttg aatatggctc ataacacccc ttgtattact
     3181 gtttatgtaa gcagacagtt ttattgttca tgaccaaaat cccttaacgt gagttttcgt
     3241 tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc
     3301 tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc
     3361 cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac
     3421 caaatactgt ccttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac
     3481 cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt
     3541 cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct
     3601 gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat
     3661 acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt
     3721 atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg
     3781 cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt
     3841 gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt
     3901 tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc cctgattctg
     3961 tggataaccg tattaccgcc tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg
     4021 agcgcagcga gtcagtgagc gaggaagcgg aagagcgcct gatgcggtat tttctcctta
     4081 cgcatctgtg cggtatttca caccgcaatg gtgcactctc agtacaatct gctctgatgc
     4141 cgcatagtta agccagtata cactccgcta tcgctacgtg actgggtcat ggctgcgccc
     4201 cgacacccgc caacacccgc tgacgcgccc tgacgggctt gtctgctccc ggcatccgct
     4261 tacagacaag ctgtgaccgt ctccgggagc tgcatgtgtc agaggttttc accgtcatca
     4321 ccgaaacgcg cgaggcagct gcggtaaagc tcatcagcgt ggtcgtgaag cgattcacag
     4381 atgtctgcct gttcatccgc gtccagctcg ttgagtttct ccagaagcgt taatgtctgg
     4441 cttctgataa agcgggccat gttaagggcg gttttttcct gtttggtcac tgatgcctcc
     4501 gtgtaagggg gatttctgtt catgggggta atgataccga tgaaacgaga gaggatgctc
     4561 acgatacggg ttactgatga tgaacatgcc cggttactgg aacgttgtga gggtaaacaa
     4621 ctggcggtat ggatgcggcg ggaccagaga aaaatcactc agggtcaatg ccagcgcttc
     4681 gttaatacag atgtaggtgt tccacagggt agccagcagc atcctgcgat gcagatccgg
     4741 aacataatgg tgcagggcgc tgacttccgc gtttccagac tttacgaaac acggaaaccg
     4801 aagaccattc atgttgttgc tcaggtcgca gacgttttgc agcagcagtc gcttcacgtt
     4861 cgctcgcgta tcggtgattc attctgctaa ccagtaaggc aaccccgcca gcctagccgg
     4921 gtcctcaacg acaggagcac gatcatgcgc acccgtgggg ccgccatgcc ggcgataatg
     4981 gcctgcttct cgccgaaacg tttggtggcg ggaccagtga cgaaggcttg agcgagggcg
     5041 tgcaagattc cgaataccgc aagcgacagg ccgatcatcg tcgcgctcca gcgaaagcgg
     5101 tcctcgccga aaatgaccca gagcgctgcc ggcacctgtc ctacgagttg catgataaag
     5161 aagacagtca taagtgcggc gacgatagtc atgccccgcg cccaccggaa ggagctgact
     5221 gggttgaagg ctctcaaggg catcggtcga gatcccggtg cctaatgagt gagctaactt
     5281 acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg
     5341 cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ccagggt
//
