LOCUS       sequence-433372-        6383 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses anti-HER2 Pert Heavy chain-SpyTag003 in mammalian cells. 
            To be paired with Pert light chain to form the anti-HER2 Pert Fab 
            (related to Pertuzumab).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6383)
  AUTHORS   Driscoll CL, Keeble AH, Howarth MR
  TITLE     SpyMask enables combinatorial assembly of bispecific binders.
  JOURNAL   Nat Commun. 2024 Mar 16;15(1):2403. doi: 10.1038/s41467-024-46599-9.
  PUBMED    38493197
REFERENCE   2  (bases 1 to 6383)
  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: "2024-03-16"; volume: "15(1):2403. doi"; pages: " 
            10.1038/s41467-024-46599-9"
COMMENT     Sequence Label: pcDNA3.1-Pert Heavy chain-SpyTag003
FEATURES             Location/Qualifiers
     source          1..6383
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      931..940
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1681..1728
                     /codon_start=1
                     /product="improved SpyTag variant that reacts with 
                     SpyCatcher003 at a rate approaching the diffusion limit 
                     (Keeble et al., 2019)"
                     /label=SpyTag003
                     /translation="RGVPHIVMVDAYKRYK"
     CDS             1732..1749
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1974..1991)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1980..2204
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2250..2678
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2337..2356)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2547..2568
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2687..2707)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2692..3021
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2872..3007
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2934..2953
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3088..3882
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3142..3161)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3752..3771
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4056..4177
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4093..4112)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4147..4166
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4226..4242)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4226..4242)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4239..4261)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4250..4266
                     /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)."
     promoter        complement(join(4274..4280,4281..4298,4299..4304))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4319..4340
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4457..4474)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4628..5216)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4708..4727)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5387..6178,6179..6247))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     6010..6029
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6248..6352)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 gtttaaacgg gccctctaga ctcgagcggc gccaccatgg ggatccttcc cagccctggg
      961 atgcctgcgc tgctctccct cgtgagcctt ctctccgtgc tgctgatggg ttgcgtagct
     1021 gaagtacagt tggtggaatc cggcggtggc ctggtgcaac ccggcgggtc gcttcgcttg
     1081 tcctgtgccg catctgggtt cactttcacg gactacacga tggactgggt tcgccaagca
     1141 ccagggaaag gtttagaatg ggttgcggat gtaaatccaa actcaggtgg ttctatctat
     1201 aatcagcgct tcaagggccg cttcacgctt agtgtggacc gttccaagaa taccttatat
     1261 cttcagatga acagtcttcg cgccgaggat acagccgttt actactgcgc acgcaatctt
     1321 ggaccatcgt tttacttcga ctactggggt caaggtacgt tggttactgt atcttcggca
     1381 agcacgaaag gaccatcggt cttccccctg gcaccctcct ccaagagcac ctctgggggc
     1441 acagcggccc tgggctgcct ggtcaaggac tacttccccg aaccggtgac ggtgtcgtgg
     1501 aactcaggcg ccctgaccag cggcgtgcac accttcccgg ctgtcctaca gtcctcagga
     1561 ctctactccc tcagcagcgt ggtgaccgtg ccctccagca gcttgggcac ccagacctac
     1621 atctgcaacg tgaatcacaa gcccagcaac accaaggtgg acaagaaagt tgggtccggg
     1681 cgtggcgtgc ctcacatcgt gatggtggac gcctacaagc gttacaaggg gcatcaccac
     1741 catcatcatt gataaaccgg taagctgagc agcggctaca aggacgtgat cctgtggttc
     1801 agcttcggcg ccagctgctt catcctgctg gccatcgtga tgggcctggt gttcatctgc
     1861 gtgaagaacg gcaacatgcg gtgcaccatc tgcatctgat gatatcgaat tccaccacac
     1921 tggactagtg gatccgagct cggtaccaag cttaagttta aaccgctgat cagcctcgac
     1981 tgtgccttct agttgccagc catctgttgt ttgcccctcc cccgtgcctt ccttgaccct
     2041 ggaaggtgcc actcccactg tcctttccta ataaaatgag gaaattgcat cgcattgtct
     2101 gagtaggtgt cattctattc tggggggtgg ggtggggcag gacagcaagg gggaggattg
     2161 ggaagacaat agcaggcatg ctggggatgc ggtgggctct atggcttctg aggcggaaag
     2221 aaccagctgg ggctctaggg ggtatcccca cgcgccctgt agcggcgcat taagcgcggc
     2281 gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc
     2341 tttcgctttc ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa
     2401 tcgggggctc cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact
     2461 tgattagggt gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt
     2521 gacgttggag tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa
     2581 ccctatctcg gtctattctt ttgatttata agggattttg ccgatttcgg cctattggtt
     2641 aaaaaatgag ctgatttaac aaaaatttaa cgcgaattaa ttctgtggaa tgtgtgtcag
     2701 ttagggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc
     2761 aattagtcag caaccaggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa
     2821 agcatgcatc tcaattagtc agcaaccata gtcccgcccc taactccgcc catcccgccc
     2881 ctaactccgc ccagttccgc ccattctccg ccccatggct gactaatttt ttttatttat
     2941 gcagaggccg aggccgcctc tgcctctgag ctattccaga agtagtgagg aggctttttt
     3001 ggaggcctag gcttttgcaa aaagctcccg ggagcttgta tatccatttt cggatctgat
     3061 caagagacag gatgaggatc gtttcgcatg attgaacaag atggattgca cgcaggttct
     3121 ccggccgctt gggtggagag gctattcggc tatgactggg cacaacagac aatcggctgc
     3181 tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc cggttctttt tgtcaagacc
     3241 gacctgtccg gtgccctgaa tgaactgcag gacgaggcag cgcggctatc gtggctggcc
     3301 acgacgggcg ttccttgcgc agctgtgctc gacgttgtca ctgaagcggg aagggactgg
     3361 ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat ctcaccttgc tcctgccgag
     3421 aaagtatcca tcatggctga tgcaatgcgg cggctgcata cgcttgatcc ggctacctgc
     3481 ccattcgacc accaagcgaa acatcgcatc gagcgagcac gtactcggat ggaagccggt
     3541 cttgtcgatc aggatgatct ggacgaagag catcaggggc tcgcgccagc cgaactgttc
     3601 gccaggctca aggcgcgcat gcccgacggc gaggatctcg tcgtgaccca tggcgatgcc
     3661 tgcttgccga atatcatggt ggaaaatggc cgcttttctg gattcatcga ctgtggccgg
     3721 ctgggtgtgg cggaccgcta tcaggacata gcgttggcta cccgtgatat tgctgaagag
     3781 cttggcggcg aatgggctga ccgcttcctc gtgctttacg gtatcgccgc tcccgattcg
     3841 cagcgcatcg ccttctatcg ccttcttgac gagttcttct gagcgggact ctggggttcg
     3901 aaatgaccga ccaagcgacg cccaacctgc catcacgaga tttcgattcc accgccgcct
     3961 tctatgaaag gttgggcttc ggaatcgttt tccgggacgc cggctggatg atcctccagc
     4021 gcggggatct catgctggag ttcttcgccc accccaactt gtttattgca gcttataatg
     4081 gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt
     4141 ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca tgtctgtata ccgtcgacct
     4201 ctagctagag cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc
     4261 tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat
     4321 gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc
     4381 tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg
     4441 ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag
     4501 cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag
     4561 gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc
     4621 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc
     4681 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc
     4741 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt
     4801 cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg
     4861 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat
     4921 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag
     4981 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt
     5041 ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc
     5101 cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta
     5161 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag
     5221 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga
     5281 ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa
     5341 gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa
     5401 tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc
     5461 ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga
     5521 taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa
     5581 gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt
     5641 gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg
     5701 ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc
     5761 aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg
     5821 gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag
     5881 cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt
     5941 actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt
     6001 caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac
     6061 gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac
     6121 ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag
     6181 caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa
     6241 tactcatact cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga
     6301 gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc
     6361 cccgaaaagt gccacctgac gtc
//
