LOCUS       sequence-369408-        6833 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Sleeping Beauty transposon system with constitutive bi-directional 
            promoter, one side expressing SpyTag003-sfCherry11 fused to PDGFR 
            through a (GGGGS)3 linker and the other side expressing BFP and 
            puromycin resistance gene.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6833)
  AUTHORS   Moghimianavval H, Patel C, Mohapatra S, Hwang SW, Kayikcioglu T, 
            Bashirzadeh Y, Liu AP, Ha T
  TITLE     Engineering Functional Membrane-Membrane Interfaces by InterSpy.
  JOURNAL   Small. 2022 May 26:e2202104. doi: 10.1002/smll.202202104.
  PUBMED    35618485
REFERENCE   2  (bases 1 to 6833)
  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: "Small. 2022
            May 26:e2202104. doi: 10.1002/smll.202202104."
COMMENT     Sequence Label: InterTag-3xL
FEATURES             Location/Qualifiers
     source          1..6833
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     154..171
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     545..564
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     polyA_signal    563..618
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     664..683
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     CDS             complement(783..1382)
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(866..886)
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     1363..1382
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     CDS             complement(1383..1439)
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             complement(1449..2147)
                     /codon_start=1
                     /product="monomeric blue fluorescent protein"
                     /label=TagBFP
                     /note="mammalian codon-optimized"
                     /translation="MSELIKENMHMKLYMEGTVDNHHFKCTSEGEGKPYEGTQTMRIKV
                     VEGGPLPFAFDILATSFLYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLTA
                     TQDTSLQDGCLIYNVKIRGVNFTSNGPVMQKKTLGWEAFTETLYPADGGLEGRNDMALK
                     LVGGSHLIANIKTTYRSKKPAKNLKMPGVYYVDYRLERIKEANNETYVEQHEVAVARYC
                     DLPSKLGHKLN"
     promoter        2779..3961
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     primer_bind     3909..3929
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     regulatory      3985..3994
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     sig_peptide     3991..4050
                     /codon_start=1
                     /product="leader sequence from mouse immunoglobulin kappa 
                     light chain"
                     /label=Ig-kappa leader
                     /translation="METDTLLLWVLLLWVPGSTG"
     CDS             4057..4104
                     /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             4219..4248
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             4252..4398
                     /codon_start=1
                     /product="transmembrane domain from platelet derived growth
                     factor receptor beta"
                     /label=PDGFR-beta TM domain
                     /translation="AVGQDTQEVIVVPHSLPFKVVVISAILALVVLTIISLIILIMLWQ
                     KKPR"
     primer_bind     complement(4447..4464)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4453..4677
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     promoter        5109..5213
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5214..5282,5283..6074)
                     /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     complement(5432..5451)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6245..6833
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6734..6753
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg
       61 ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgcctt tgagtgagct
      121 gataccgctc gccgcagccg aacgaccgag cgcagcgagt cagtgagcga ggaagcggat
      181 ccctatacag ttgaagtcgg aagtttacat acaccttagc caaatacatt taaactcact
      241 ttttcacaat tcctgacatt taatcctagt aaaaattccc tgtcttaggt cagttaggat
      301 caccacttta ttttaagaat gtgaaatatc agaataatag tagagagaat gattcatttc
      361 agcttttatt tctttcatca cattcccagt gggtcagaag tttacataca ctcgactcct
      421 ctgcagaatg cggcgatgtt tcggtaaggg gtcccgagta gctagttcat ggcagccagc
      481 atatggcata tgttgccaaa ctctaaacca aatactcatt ctgatgtttt aaatgatttg
      541 ccctcccata tgtccttccg agtgagagac acaaaaaatt ccaacacact attgcaatga
      601 aaataaattt cctttattag ccagaagtca gatgctcaag gggcttcatg atgtccccat
      661 aatttttggc agagggaaaa agatctcagt ggtatttgtg agccagggca ttggccacac
      721 cagccaccac cttctgatag gcagcctgca cctgaggagt gaattcacga caggccttcg
      781 aatcaggcac cgggcttgcg ggtcatgcac caggtgcgcg gtccttcggg cacctcgacg
      841 tcggcggtga cggtgaagcc gagccgctcg tagaagggga ggttgcgggg cgcggaggtc
      901 tccaggaagg cgggcacccc ggcgcgctcg gccgcctcca ctccggggag cacgacggcg
      961 ctgcccagac ccttgccctg gtggtcgggc gagacgccga cggtggccag gaaccacgcg
     1021 ggctccttgg gccggtgcgg cgccaggagg ccttccatct gttgctgcgc ggccagccgg
     1081 gaaccgctca actcggccat gcgcgggccg atctcggcga acaccgcccc cgcttcgacg
     1141 ctctccggcg tggtccagac cgccaccgcg gcgccgtcgt ccgcgaccca caccttgccg
     1201 atgtcgagcc cgacgcgcgt gaggaagagt tcttgcagct cggtgacccg ctcgatgtgg
     1261 cggtccggat cgacggtgtg gcgcgtggcg gggtagtcgg cgaacgcggc ggcgagggtg
     1321 cgtacggccc tggggacgtc gtcgcgggtg gcgaggcgca ccgtgggctt gtactcggtc
     1381 atggggccag gattctcctc gacgtcacca gcctgcttca gcaggctgaa gttagtagct
     1441 ccactgccgt tcagcttgtg ccccagtttg ctagggaggt cgcagtatct ggccactgcc
     1501 acctcgtgct gctcgacgta ggtctcgttg ttggcctcct tgattctttc cagtctgtag
     1561 tccacatagt agacgccagg catcttgagg ttcttagcgg gtttcttgga tctatatgtg
     1621 gtcttgatgt ttgcgatcag atggctcccg cccacgagct tcagggccat gtcgtttctg
     1681 ccttccaggc cgccgtcagc ggggtacagc gtctcggtga aggcctccca gccgagtgtt
     1741 ttcttctgca tcacagggcc gttggatgtg aagttcaccc ctctgatctt gacgttgtag
     1801 atgaggcagc cgtcctggag gctggtgtcc tgggtagcgg tcagcacgcc cccgtcttcg
     1861 tatgtggtga ctctctccca tgtgaagccc tcagggaagg actgcttgaa gaagtcgggg
     1921 atgccctggg tgtggttgat gaaggtcttg ctgccgtaga ggaagctagt agccaggatg
     1981 tcgaaggcga aggggagagg gccgccctcg accaccttga ttctcatggt ctgggtgccc
     2041 tcgtagggct tgccttcgcc ctcggatgtg cacttgaagt gatggttgtc cacggtgccc
     2101 tccatgtaca gcttcatgtg catgttctcc ttaatcagct cgctcatggt ggcctcaggt
     2161 gcagaggttt ctacagggaa agagagaaag gctacagtta gtttgctgtg atgatgctaa
     2221 gtgccgaggt cagatgtcta tggattgtac acggaatccc aacgcaggaa ccaaaaaaca
     2281 ctcacccaaa aaaatggcgc taaggccgag agaaaggcga agttccgtct acggctatat
     2341 accttgaaac aggggcggca gaagtgacgc aacgcggttg cgcggacggc ttcctgtgtt
     2401 ttcattggtc gaagcgttct gcgcgtgcgc agagattccc cagggaaggc gggccctctt
     2461 ccgaatgtcc aatccggaca ttcttccggt tggacctccg cggtcagcgc tgtgcccggc
     2521 gagagatcac gtggggcgcg gaggcggtgc tgctggggca cggccgtcca gcctcggcgg
     2581 ccatattttt gaggggctgt tcatctcgtt cacacgctct gtccgccatg tttgtgagtg
     2641 gaagcgccat taccccttca agcgactgaa ggctgcaggg cctctggtgg cccgcatggg
     2701 gagaccagac ccgccaggcc cgcctttccg cactcagtcc gggcttactt tattttgtga
     2761 gacagggtct cgcctagagg ctccggtgcc cgtcagtggg cagagcgcac atcgcccaca
     2821 gtccccgaga agttgggggg aggggtcggc aattgaaccg gtgcctagag aaggtggcgc
     2881 ggggtaaact gggaaagtga tgtcgtgtac tggctccgcc tttttcccga gggtggggga
     2941 gaaccgtata taagtgcagt agtcgccgtg aacgttcttt ttcgcaacgg gtttgccgcc
     3001 agaacacagg taagtgccgt gtgtggttcc cgcgggcctg gcctctttac gggttatggc
     3061 ccttgcgtgc cttgaattac ttccacgccc ctggctgcag tacgtgattc ttgatcccga
     3121 gcttcgggtt ggaagtgggt gggagagttc gaggccttgc gcttaaggag ccccttcgcc
     3181 tcgtgcttga gttgaggcct ggcttgggcg ctggggccgc cgcgtgcgaa tctggtggca
     3241 ccttcgcgcc tgtctcgctg ctttcgataa gtctctagcc atttaaaatt tttgatgacc
     3301 tgctgcgacg ctttttttct ggcaagatag tcttgtaaat gcgggccaag atctgcacac
     3361 tggtatttcg gtttttgggg ccgcgggcgg cgacggggcc cgtgcgtccc agcgcacatg
     3421 ttcggcgagg cggggcctgc gagcgcggcc accgagaatc ggacgggggt agtctcaagc
     3481 tggccggcct gctctggtgc ctggcctcgc gccgccgtgt atcgccccgc cctgggcggc
     3541 aaggctggcc cggtcggcac cagttgcgtg agcggaaaga tggccgcttc ccggccctgc
     3601 tgcagggagc tcaaaatgga ggacgcggcg ctcgggagag cgggcgggtg agtcacccac
     3661 acaaaggaaa agggcctttc cgtcctcagc cgtcgcttca tgtgactcca cggagtaccg
     3721 ggcgccgtcc aggcacctcg attagttctc gagcttttgg agtacgtcgt ctttaggttg
     3781 gggggagggg ttttatgcga tggagtttcc ccatactgag tgggtggaga ctgaagttag
     3841 gccagcttgg cacttgatgt aattctcctt ggaatttgcc ctttttgagt ttggatcttg
     3901 gttcattctc aagcctcaga cagtggttca aagttttttt cttccattta aggtgtcgtg
     3961 aaaactaccc caagctggcc tctggccacc atggagacag acacactcct gctatgggta
     4021 ctgctgctct gggttccagg ttccactggt gacggacgtg gcgttcctca tattgttatg
     4081 gtggacgcct acaaacgcta taaatcggga ggtggttcag gctacaccat cgtggagcag
     4141 tacgagagag ccgaggccag acacagcacc gcgggaggcg gtggatcggg aggcggtgga
     4201 tcgggaggcg gtggatcgga acaaaaactc atctcagaag aggatctgaa tgctgtgggc
     4261 caggacacgc aggaggtcat cgtggtgcca cactccttgc cctttaaggt ggtggtgatc
     4321 tcagccatcc tggccctggt ggtgctcacc atcatctccc ttatcatcct catcatgctt
     4381 tggcagaaga agccacgtta ggtcaggcca agcttgggga tcaattctct agagctcgct
     4441 gatcagcctc gactgtgcct tctagttgcc agccatctgt tgtttgcccc tcccccgtgc
     4501 cttccttgac cctggaaggt gccactccca ctgtcctttc ctaataaaat gaggaaattg
     4561 catcgcattg tctgagtagg tgtcattcta ttctgggggg tggggtgggg caggacagca
     4621 agggggagga ttgggaagac aatagcaggc atgctgggga tgcggtgggc tctatggctt
     4681 ctgaggcgga aagaaccagc tgggcctact agatagcgga ccccttaccg aaacatcgcc
     4741 gcattctgca gaggagtcga acttagtgta tgtaaacttc tgacccactg gaattgtgat
     4801 acagtgaatt ataagtgaaa taatctgtct gtaaacaatt gttggaaaaa tgacttgtgt
     4861 catgcacaaa gtagatgtcc taactgactt gccaaaacta ttgtttgtta acaagaaatt
     4921 tgtggagtag ttgaaaaacg agttttaatg actccaactt aagtgtatgt aaacttccga
     4981 cttcaactgt atagggatcc aattttaaca aaatattaac gcttacaatt tcctgatgcg
     5041 gtattttctc cttacgcatc tgtgcggtat ttcacaccgc atcaggtggc acttttcggg
     5101 gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc
     5161 tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta
     5221 ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg
     5281 ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg
     5341 gttacatcga actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac
     5401 gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg
     5461 acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt
     5521 actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg
     5581 ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac
     5641 cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt
     5701 gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgtag
     5761 caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc
     5821 aacaattaat agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc
     5881 ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta
     5941 tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg
     6001 ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga
     6061 ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac
     6121 ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa
     6181 tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat
     6241 cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc
     6301 taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg
     6361 gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc
     6421 acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg
     6481 ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg
     6541 ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa
     6601 cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg
     6661 aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga
     6721 gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct
     6781 gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaa
//
