LOCUS       sequence-207217-        6559 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid for recombinant bacterial expression of SnoopLigase, a 
            protein that promotes the formation of an isopeptide bond between 2 
            peptide tags (SnoopTagJr and DogTag), with N-terminal HaloTag7.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6559)
  AUTHORS   Buldun CM, Jean JX, Bedford MR, Howarth M
  TITLE     SnoopLigase Catalyzes Peptide-Peptide Locking and Enables 
            Solid-Phase Conjugate Isolation.
  JOURNAL   J Am Chem Soc. 2018 Feb 28;140(8):3008-3018. doi: 
            10.1021/jacs.7b13237. Epub 2018 Feb 14.
  PUBMED    29402082
REFERENCE   2  (bases 1 to 6559)
  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/jacs.7b13237"; journalName: "J Am Chem Soc"; date: 
            "2018-02-28- 28"; volume: "140"; issue: "8"; pages: "3008-3018"
COMMENT     Sequence Label: pET28a-HaloTag7-SnoopLigase
FEATURES             Location/Qualifiers
     source          1..6559
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     126..145
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     229..248
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        229..247
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    248..272
                     /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             287..309
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             328..345
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             355..1239
                     /codon_start=1
                     /product="modified bacterial dehalogenase that forms 
                     covalent bonds with chloroalkane derivatives"
                     /label=HaloTag(R)
                     /translation="EIGTGFPFDPHYVEVLGERMHYVDVGPRDGTPVLFLHGNPTSSYV
                     WRNIIPHVAPTHRCIAPDLIGMGKSDKPDLGYFFDDHVRFMDAFIEALGLEEVVLVIHD
                     WGSALGFHWAKRNPERVKGIAFMEFIRPIPTWDEWPEFARETFQAFRTTDVGRKLIIDQ
                     NVFIEGTLPMGVVRPLTEVEMDHYREPFLNPVDREPLWRFPNELPIAGEPANIVALVEE
                     YMDWLHQSPVPKLLFWGTPGVLIPPAEAARLAKSLPNCKAVDIGPGLNLLQEDNPDLIG
                     SEIARWLSTLEISG"
     primer_bind     complement(1721..1739)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1735..1782
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1819..2274
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1906..1925)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2116..2137
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(2366..3181)
                     /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(2366..3181)
                     /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     3089..3108
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     3173..3192
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      3303..3891
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3792..3811
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4045..4062
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4077..4216
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4142..4161)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4302..4324
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4318..4509)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(5318..6400)
                     /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     6362..6381
                     /label=LacI-R
                     /note="LacI, reverse primer"
ORIGIN
        1 gccgcaagga atggtgcatg caaggagatg gcgcccaaca gtcccccggc cacggggcct
       61 gccaccatac ccacgccgaa acaagcgctc atgagcccga agtggcgagc ccgatcttcc
      121 ccatcggtga tgtcggcgat ataggcgcca gcaaccgcac ctgtggcgcc ggtgatgccg
      181 gccacgatgc gtccggcgta gaggatcgag atctcgatcc cgcgaaatta atacgactca
      241 ctatagggga attgtgagcg gataacaatt cccctctaga aataattttg tttaacttta
      301 agaaggagat ataccatggg cagcagccat catcatcatc atcacagcag cggcgaaatc
      361 ggtactggct ttccattcga cccccattat gtggaagtcc tgggcgagcg catgcactac
      421 gtcgatgttg gtccgcgcga tggcacccct gtgctgttcc tgcacggtaa cccgacctcc
      481 tcctacgtgt ggcgcaacat catcccgcat gttgcaccga cccatcgctg cattgctcca
      541 gacctgatcg gtatgggcaa atccgacaaa ccagacctgg gttatttctt cgacgaccac
      601 gtccgcttca tggatgcctt catcgaagcc ctgggtctgg aagaggtcgt cctggtcatt
      661 cacgactggg gctccgctct gggtttccac tgggccaagc gcaatccaga gcgcgtcaaa
      721 ggtattgcat ttatggagtt catccgccct atcccgacct gggacgaatg gccagaattt
      781 gcccgcgaga ccttccaggc cttccgcacc accgacgtcg gccgcaagct gatcatcgat
      841 cagaacgttt ttatcgaggg tacgctgccg atgggtgtcg tccgcccgct gactgaagtc
      901 gagatggacc attaccgcga gccgttcctg aatcctgttg accgcgagcc actgtggcgc
      961 ttcccaaacg agctgccaat cgccggtgag ccagcgaaca tcgtcgcgct ggtcgaagaa
     1021 tacatggact ggctgcacca gtcccctgtc ccgaagctgc tgttctgggg caccccaggc
     1081 gttctgatcc caccggccga agccgctcgc ctggccaaaa gcctgcctaa ctgcaaggct
     1141 gtggacatcg gcccgggtct gaatctgctg caagaagaca acccggacct gatcggcagc
     1201 gagatcgcgc gctggctgtc tactctggag atttccggtg gtagcggtgg gtctggtagt
     1261 ggtgtgaata agaacgataa aaagccgctg cgtggtgccg tgtttagcct gcagaaacag
     1321 catcccgact atcccgatat ctatggcgcg attgatcaga atgggaccta tcaaaatgtg
     1381 cgtaccggcg aagatggtaa actgaccttt aagaatctga gcgatggcaa atatcgcctg
     1441 tttgaaaata gcgaaccccc gggctataaa ccggtgcaga ataagccgat tgtggcgttt
     1501 cagattgtga atggcgaagt gcgtgatgtg accagcattg tgccgccggg tgtgccggct
     1561 acatatgaat ttacctaata aaacggtaaa cattatatca ccaatgaacc gataccgccg
     1621 aaataataaa agcttgcggc cgcactcgag cactaccacc accaccactg agatccggct
     1681 gctaacaaag cccgaaagga agctgagttg gctgctgcca ccgctgagca ataactagca
     1741 taaccccttg gggcctctaa acgggtcttg aggggttttt tgctgaaagg aggaactata
     1801 tccggattgg cgaatgggac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg
     1861 ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct
     1921 tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc
     1981 ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt gattagggtg
     2041 atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt
     2101 ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac cctatctcgg
     2161 tctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta aaaaatgagc
     2221 tgatttaaca aaaatttaac gcgaatttta acaaaatatt aacgcttaca atttaggtgg
     2281 cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa
     2341 tatgtatccg ctcatgaatt aattcttaga aaaactcatc gagcatcaaa tgaaactgca
     2401 atttattcat atcaggatta tcaataccat atttttgaaa aagccgtttc tgtaatgaag
     2461 gagaaaactc accgaggcag ttccatagga tggcaagatc ctggtatcgg tctgcgattc
     2521 cgactcgtcc aacatcaata caacctatta atttcccctc gtcaaaaata aggttatcaa
     2581 gtgagaaatc accatgagtg acgactgaat ccggtgagaa tggcaaaagt ttatgcattt
     2641 ctttccagac ttgttcaaca ggccagccat tacgctcgtc atcaaaatca ctcgcatcaa
     2701 ccaaaccgtt attcattcgt gattgcgcct gagcgagacg aaatacgcga tcgctgttaa
     2761 aaggacaatt acaaacagga atcgaatgca accggcgcag gaacactgcc agcgcatcaa
     2821 caatattttc acctgaatca ggatattctt ctaatacctg gaatgctgtt ttcccgggga
     2881 tcgcagtggt gagtaaccat gcatcatcag gagtacggat aaaatgcttg atggtcggaa
     2941 gaggcataaa ttccgtcagc cagtttagtc tgaccatctc atctgtaaca tcattggcaa
     3001 cgctaccttt gccatgtttc agaaacaact ctggcgcatc gggcttccca tacaatcgat
     3061 agattgtcgc acctgattgc ccgacattat cgcgagccca tttataccca tataaatcag
     3121 catccatgtt ggaatttaat cgcggcctag agcaagacgt ttcccgttga atatggctca
     3181 taacacccct tgtattactg tttatgtaag cagacagttt tattgttcat gaccaaaatc
     3241 ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct
     3301 tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta
     3361 ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc
     3421 ttcagcagag cgcagatacc aaatactgtc cttctagtgt agccgtagtt aggccaccac
     3481 ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct
     3541 gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat
     3601 aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg
     3661 acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa
     3721 gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg
     3781 gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga
     3841 cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc
     3901 aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat gttctttcct
     3961 gcgttatccc ctgattctgt ggataaccgt attaccgcct ttgagtgagc tgataccgct
     4021 cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg aggaagcgga agagcgcctg
     4081 atgcggtatt ttctccttac gcatctgtgc ggtatttcac accgcaatgg tgcactctca
     4141 gtacaatctg ctctgatgcc gcatagttaa gccagtatac actccgctat cgctacgtga
     4201 ctgggtcatg gctgcgcccc gacacccgcc aacacccgct gacgcgccct gacgggcttg
     4261 tctgctcccg gcatccgctt acagacaagc tgtgaccgtc tccgggagct gcatgtgtca
     4321 gaggttttca ccgtcatcac cgaaacgcgc gaggcagctg cggtaaagct catcagcgtg
     4381 gtcgtgaagc gattcacaga tgtctgcctg ttcatccgcg tccagctcgt tgagtttctc
     4441 cagaagcgtt aatgtctggc ttctgataaa gcgggccatg ttaagggcgg ttttttcctg
     4501 tttggtcact gatgcctccg tgtaaggggg atttctgttc atgggggtaa tgataccgat
     4561 gaaacgagag aggatgctca cgatacgggt tactgatgat gaacatgccc ggttactgga
     4621 acgttgtgag ggtaaacaac tggcggtatg gatgcggcgg gaccagagaa aaatcactca
     4681 gggtcaatgc cagcgcttcg ttaatacaga tgtaggtgtt ccacagggta gccagcagca
     4741 tcctgcgatg cagatccgga acataatggt gcagggcgct gacttccgcg tttccagact
     4801 ttacgaaaca cggaaaccga agaccattca tgttgttgct caggtcgcag acgttttgca
     4861 gcagcagtcg cttcacgttc gctcgcgtat cggtgattca ttctgctaac cagtaaggca
     4921 accccgccag cctagccggg tcctcaacga caggagcacg atcatgcgca cccgtggggc
     4981 cgccatgccg gcgataatgg cctgcttctc gccgaaacgt ttggtggcgg gaccagtgac
     5041 gaaggcttga gcgagggcgt gcaagattcc gaataccgca agcgacaggc cgatcatcgt
     5101 cgcgctccag cgaaagcggt cctcgccgaa aatgacccag agcgctgccg gcacctgtcc
     5161 tacgagttgc atgataaaga agacagtcat aagtgcggcg acgatagtca tgccccgcgc
     5221 ccaccggaag gagctgactg ggttgaaggc tctcaagggc atcggtcgag atcccggtgc
     5281 ctaatgagtg agctaactta cattaattgc gttgcgctca ctgcccgctt tccagtcggg
     5341 aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg
     5401 tattgggcgc cagggtggtt tttcttttca ccagtgagac gggcaacagc tgattgccct
     5461 tcaccgcctg gccctgagag agttgcagca agcggtccac gctggtttgc cccagcaggc
     5521 gaaaatcctg tttgatggtg gttaacggcg ggatataaca tgagctgtct tcggtatcgt
     5581 cgtatcccac taccgagata tccgcaccaa cgcgcagccc ggactcggta atggcgcgca
     5641 ttgcgcccag cgccatctga tcgttggcaa ccagcatcgc agtgggaacg atgccctcat
     5701 tcagcatttg catggtttgt tgaaaaccgg acatggcact ccagtcgcct tcccgttccg
     5761 ctatcggctg aatttgattg cgagtgagat atttatgcca gccagccaga cgcagacgcg
     5821 ccgagacaga acttaatggg cccgctaaca gcgcgatttg ctggtgaccc aatgcgacca
     5881 gatgctccac gcccagtcgc gtaccgtctt catgggagaa aataatactg ttgatgggtg
     5941 tctggtcaga gacatcaaga aataacgccg gaacattagt gcaggcagct tccacagcaa
     6001 tggcatcctg gtcatccagc ggatagttaa tgatcagccc actgacgcgt tgcgcgagaa
     6061 gattgtgcac cgccgcttta caggcttcga cgccgcttcg ttctaccatc gacaccacca
     6121 cgctggcacc cagttgatcg gcgcgagatt taatcgccgc gacaatttgc gacggcgcgt
     6181 gcagggccag actggaggtg gcaacgccaa tcagcaacga ctgtttgccc gccagttgtt
     6241 gtgccacgcg gttgggaatg taattcagct ccgccatcgc cgcttccact ttttcccgcg
     6301 ttttcgcaga aacgtggctg gcctggttca ccacgcggga aacggtctga taagagacac
     6361 cggcatactc tgcgacatcg tataacgtta ctggtttcac attcgccacc tctgacttga
     6421 ctctcttccg ggcgctatca tgccataccg cgaaaggttt tgcgccattc gatggtgtcc
     6481 gggatctcga cgctctccct tatgcgactc ctgcattagg aagcagccca gtagtaggtt
     6541 gaggccgttg agcaccgcc
//
