LOCUS       sequence-369407-        6818 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)2 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 6818)
  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 6818)
  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-2xL
FEATURES             Location/Qualifiers
     source          1..6818
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     743..760
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     1134..1153
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     polyA_signal    1152..1207
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     1253..1272
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     CDS             complement(1372..1971)
                     /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(1455..1475)
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     1952..1971
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     CDS             complement(1972..2028)
                     /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(2038..2736)
                     /codon_start=1
                     /product="monomeric blue fluorescent protein"
                     /label=TagBFP
                     /note="mammalian codon-optimized"
                     /translation="MSELIKENMHMKLYMEGTVDNHHFKCTSEGEGKPYEGTQTMRIKV
                     VEGGPLPFAFDILATSFLYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLTA
                     TQDTSLQDGCLIYNVKIRGVNFTSNGPVMQKKTLGWEAFTETLYPADGGLEGRNDMALK
                     LVGGSHLIANIKTTYRSKKPAKNLKMPGVYYVDYRLERIKEANNETYVEQHEVAVARYC
                     DLPSKLGHKLN"
     promoter        3368..4550
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     primer_bind     4498..4518
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     regulatory      4574..4583
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     sig_peptide     4580..4639
                     /codon_start=1
                     /product="leader sequence from mouse immunoglobulin kappa 
                     light chain"
                     /label=Ig-kappa leader
                     /translation="METDTLLLWVLLLWVPGSTG"
     CDS             4646..4693
                     /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             4793..4822
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             4826..4972
                     /codon_start=1
                     /product="transmembrane domain from platelet derived growth
                     factor receptor beta"
                     /label=PDGFR-beta TM domain
                     /translation="AVGQDTQEVIVVPHSLPFKVVVISAILALVVLTIISLIILIMLWQ
                     KKPR"
     primer_bind     complement(5021..5038)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5027..5251
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     promoter        5683..5787
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5788..5856,5857..6648)
                     /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(6006..6025)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
      661 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
      721 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggatc cctatacagt
      781 tgaagtcgga agtttacata caccttagcc aaatacattt aaactcactt tttcacaatt
      841 cctgacattt aatcctagta aaaattccct gtcttaggtc agttaggatc accactttat
      901 tttaagaatg tgaaatatca gaataatagt agagagaatg attcatttca gcttttattt
      961 ctttcatcac attcccagtg ggtcagaagt ttacatacac tcgactcctc tgcagaatgc
     1021 ggcgatgttt cggtaagggg tcccgagtag ctagttcatg gcagccagca tatggcatat
     1081 gttgccaaac tctaaaccaa atactcattc tgatgtttta aatgatttgc cctcccatat
     1141 gtccttccga gtgagagaca caaaaaattc caacacacta ttgcaatgaa aataaatttc
     1201 ctttattagc cagaagtcag atgctcaagg ggcttcatga tgtccccata atttttggca
     1261 gagggaaaaa gatctcagtg gtatttgtga gccagggcat tggccacacc agccaccacc
     1321 ttctgatagg cagcctgcac ctgaggagtg aattcacgac aggccttcga atcaggcacc
     1381 gggcttgcgg gtcatgcacc aggtgcgcgg tccttcgggc acctcgacgt cggcggtgac
     1441 ggtgaagccg agccgctcgt agaaggggag gttgcggggc gcggaggtct ccaggaaggc
     1501 gggcaccccg gcgcgctcgg ccgcctccac tccggggagc acgacggcgc tgcccagacc
     1561 cttgccctgg tggtcgggcg agacgccgac ggtggccagg aaccacgcgg gctccttggg
     1621 ccggtgcggc gccaggaggc cttccatctg ttgctgcgcg gccagccggg aaccgctcaa
     1681 ctcggccatg cgcgggccga tctcggcgaa caccgccccc gcttcgacgc tctccggcgt
     1741 ggtccagacc gccaccgcgg cgccgtcgtc cgcgacccac accttgccga tgtcgagccc
     1801 gacgcgcgtg aggaagagtt cttgcagctc ggtgacccgc tcgatgtggc ggtccggatc
     1861 gacggtgtgg cgcgtggcgg ggtagtcggc gaacgcggcg gcgagggtgc gtacggccct
     1921 ggggacgtcg tcgcgggtgg cgaggcgcac cgtgggcttg tactcggtca tggggccagg
     1981 attctcctcg acgtcaccag cctgcttcag caggctgaag ttagtagctc cactgccgtt
     2041 cagcttgtgc cccagtttgc tagggaggtc gcagtatctg gccactgcca cctcgtgctg
     2101 ctcgacgtag gtctcgttgt tggcctcctt gattctttcc agtctgtagt ccacatagta
     2161 gacgccaggc atcttgaggt tcttagcggg tttcttggat ctatatgtgg tcttgatgtt
     2221 tgcgatcaga tggctcccgc ccacgagctt cagggccatg tcgtttctgc cttccaggcc
     2281 gccgtcagcg gggtacagcg tctcggtgaa ggcctcccag ccgagtgttt tcttctgcat
     2341 cacagggccg ttggatgtga agttcacccc tctgatcttg acgttgtaga tgaggcagcc
     2401 gtcctggagg ctggtgtcct gggtagcggt cagcacgccc ccgtcttcgt atgtggtgac
     2461 tctctcccat gtgaagccct cagggaagga ctgcttgaag aagtcgggga tgccctgggt
     2521 gtggttgatg aaggtcttgc tgccgtagag gaagctagta gccaggatgt cgaaggcgaa
     2581 ggggagaggg ccgccctcga ccaccttgat tctcatggtc tgggtgccct cgtagggctt
     2641 gccttcgccc tcggatgtgc acttgaagtg atggttgtcc acggtgccct ccatgtacag
     2701 cttcatgtgc atgttctcct taatcagctc gctcatggtg gcctcaggtg cagaggtttc
     2761 tacagggaaa gagagaaagg ctacagttag tttgctgtga tgatgctaag tgccgaggtc
     2821 agatgtctat ggattgtaca cggaatccca acgcaggaac caaaaaacac tcacccaaaa
     2881 aaatggcgct aaggccgaga gaaaggcgaa gttccgtcta cggctatata ccttgaaaca
     2941 ggggcggcag aagtgacgca acgcggttgc gcggacggct tcctgtgttt tcattggtcg
     3001 aagcgttctg cgcgtgcgca gagattcccc agggaaggcg ggccctcttc cgaatgtcca
     3061 atccggacat tcttccggtt ggacctccgc ggtcagcgct gtgcccggcg agagatcacg
     3121 tggggcgcgg aggcggtgct gctggggcac ggccgtccag cctcggcggc catatttttg
     3181 aggggctgtt catctcgttc acacgctctg tccgccatgt ttgtgagtgg aagcgccatt
     3241 accccttcaa gcgactgaag gctgcagggc ctctggtggc ccgcatgggg agaccagacc
     3301 cgccaggccc gcctttccgc actcagtccg ggcttacttt attttgtgag acagggtctc
     3361 gcctagaggc tccggtgccc gtcagtgggc agagcgcaca tcgcccacag tccccgagaa
     3421 gttgggggga ggggtcggca attgaaccgg tgcctagaga aggtggcgcg gggtaaactg
     3481 ggaaagtgat gtcgtgtact ggctccgcct ttttcccgag ggtgggggag aaccgtatat
     3541 aagtgcagta gtcgccgtga acgttctttt tcgcaacggg tttgccgcca gaacacaggt
     3601 aagtgccgtg tgtggttccc gcgggcctgg cctctttacg ggttatggcc cttgcgtgcc
     3661 ttgaattact tccacgcccc tggctgcagt acgtgattct tgatcccgag cttcgggttg
     3721 gaagtgggtg ggagagttcg aggccttgcg cttaaggagc cccttcgcct cgtgcttgag
     3781 ttgaggcctg gcttgggcgc tggggccgcc gcgtgcgaat ctggtggcac cttcgcgcct
     3841 gtctcgctgc tttcgataag tctctagcca tttaaaattt ttgatgacct gctgcgacgc
     3901 tttttttctg gcaagatagt cttgtaaatg cgggccaaga tctgcacact ggtatttcgg
     3961 tttttggggc cgcgggcggc gacggggccc gtgcgtccca gcgcacatgt tcggcgaggc
     4021 ggggcctgcg agcgcggcca ccgagaatcg gacgggggta gtctcaagct ggccggcctg
     4081 ctctggtgcc tggcctcgcg ccgccgtgta tcgccccgcc ctgggcggca aggctggccc
     4141 ggtcggcacc agttgcgtga gcggaaagat ggccgcttcc cggccctgct gcagggagct
     4201 caaaatggag gacgcggcgc tcgggagagc gggcgggtga gtcacccaca caaaggaaaa
     4261 gggcctttcc gtcctcagcc gtcgcttcat gtgactccac ggagtaccgg gcgccgtcca
     4321 ggcacctcga ttagttctcg agcttttgga gtacgtcgtc tttaggttgg ggggaggggt
     4381 tttatgcgat ggagtttccc catactgagt gggtggagac tgaagttagg ccagcttggc
     4441 acttgatgta attctccttg gaatttgccc tttttgagtt tggatcttgg ttcattctca
     4501 agcctcagac agtggttcaa agtttttttc ttccatttaa ggtgtcgtga aaactacccc
     4561 aagctggcct ctggccacca tggagacaga cacactcctg ctatgggtac tgctgctctg
     4621 ggttccaggt tccactggtg acggacgtgg cgttcctcat attgttatgg tggacgccta
     4681 caaacgctat aaatcgggag gtggttcagg ctacaccatc gtggagcagt acgagagagc
     4741 cgaggccaga cacagcaccg cgggaggcgg tggatcggga ggcggtggat cggaacaaaa
     4801 actcatctca gaagaggatc tgaatgctgt gggccaggac acgcaggagg tcatcgtggt
     4861 gccacactcc ttgcccttta aggtggtggt gatctcagcc atcctggccc tggtggtgct
     4921 caccatcatc tcccttatca tcctcatcat gctttggcag aagaagccac gttaggtcag
     4981 gccaagcttg gggatcaatt ctctagagct cgctgatcag cctcgactgt gccttctagt
     5041 tgccagccat ctgttgtttg cccctccccc gtgccttcct tgaccctgga aggtgccact
     5101 cccactgtcc tttcctaata aaatgaggaa attgcatcgc attgtctgag taggtgtcat
     5161 tctattctgg ggggtggggt ggggcaggac agcaaggggg aggattggga agacaatagc
     5221 aggcatgctg gggatgcggt gggctctatg gcttctgagg cggaaagaac cagctgggcc
     5281 tactagatag cggacccctt accgaaacat cgccgcattc tgcagaggag tcgaacttag
     5341 tgtatgtaaa cttctgaccc actggaattg tgatacagtg aattataagt gaaataatct
     5401 gtctgtaaac aattgttgga aaaatgactt gtgtcatgca caaagtagat gtcctaactg
     5461 acttgccaaa actattgttt gttaacaaga aatttgtgga gtagttgaaa aacgagtttt
     5521 aatgactcca acttaagtgt atgtaaactt ccgacttcaa ctgtataggg atccaatttt
     5581 aacaaaatat taacgcttac aatttcctga tgcggtattt tctccttacg catctgtgcg
     5641 gtatttcaca ccgcatcagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt
     5701 ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg
     5761 cttcaataat attgaaaaag gaagagtatg agtattcaac atttccgtgt cgcccttatt
     5821 cccttttttg cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta
     5881 aaagatgctg aagatcagtt gggtgcacga gtgggttaca tcgaactgga tctcaacagc
     5941 ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc caatgatgag cacttttaaa
     6001 gttctgctat gtggcgcggt attatcccgt attgacgccg ggcaagagca actcggtcgc
     6061 cgcatacact attctcagaa tgacttggtt gagtactcac cagtcacaga aaagcatctt
     6121 acggatggca tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact
     6181 gcggccaact tacttctgac aacgatcgga ggaccgaagg agctaaccgc ttttttgcac
     6241 aacatggggg atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata
     6301 ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta
     6361 ttaactggcg aactacttac tctagcttcc cggcaacaat taatagactg gatggaggcg
     6421 gataaagttg caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat
     6481 aaatctggag ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt
     6541 aagccctccc gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga
     6601 aatagacaga tcgctgagat aggtgcctca ctgattaagc attggtaact gtcagaccaa
     6661 gtttactcat atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag
     6721 gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac
     6781 tgagcgtcag accccgtaga aaagatcaaa ggatcttc
//
