LOCUS       sequence-369406-        6803 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 single GGGGS 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 6803)
  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 6803)
  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-1xL
FEATURES             Location/Qualifiers
     source          1..6803
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     257..276
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     polyA_signal    275..330
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     376..395
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     CDS             complement(495..1094)
                     /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(578..598)
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     1075..1094
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     CDS             complement(1095..1151)
                     /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(1161..1859)
                     /codon_start=1
                     /product="monomeric blue fluorescent protein"
                     /label=TagBFP
                     /note="mammalian codon-optimized"
                     /translation="MSELIKENMHMKLYMEGTVDNHHFKCTSEGEGKPYEGTQTMRIKV
                     VEGGPLPFAFDILATSFLYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLTA
                     TQDTSLQDGCLIYNVKIRGVNFTSNGPVMQKKTLGWEAFTETLYPADGGLEGRNDMALK
                     LVGGSHLIANIKTTYRSKKPAKNLKMPGVYYVDYRLERIKEANNETYVEQHEVAVARYC
                     DLPSKLGHKLN"
     promoter        2491..3673
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     primer_bind     3621..3641
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     regulatory      3697..3706
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     sig_peptide     3703..3762
                     /codon_start=1
                     /product="leader sequence from mouse immunoglobulin kappa 
                     light chain"
                     /label=Ig-kappa leader
                     /translation="METDTLLLWVLLLWVPGSTG"
     CDS             3769..3816
                     /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             3901..3930
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             3934..4080
                     /codon_start=1
                     /product="transmembrane domain from platelet derived growth
                     factor receptor beta"
                     /label=PDGFR-beta TM domain
                     /translation="AVGQDTQEVIVVPHSLPFKVVVISAILALVVLTIISLIILIMLWQ
                     KKPR"
     primer_bind     complement(4129..4146)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4135..4359
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     promoter        4791..4895
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4896..4964,4965..5756)
                     /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(5114..5133)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5927..6515
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6416..6435
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6669..6686
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 gtcagttagg atcaccactt tattttaaga atgtgaaata tcagaataat agtagagaga
       61 atgattcatt tcagctttta tttctttcat cacattccca gtgggtcaga agtttacata
      121 cactcgactc ctctgcagaa tgcggcgatg tttcggtaag gggtcccgag tagctagttc
      181 atggcagcca gcatatggca tatgttgcca aactctaaac caaatactca ttctgatgtt
      241 ttaaatgatt tgccctccca tatgtccttc cgagtgagag acacaaaaaa ttccaacaca
      301 ctattgcaat gaaaataaat ttcctttatt agccagaagt cagatgctca aggggcttca
      361 tgatgtcccc ataatttttg gcagagggaa aaagatctca gtggtatttg tgagccaggg
      421 cattggccac accagccacc accttctgat aggcagcctg cacctgagga gtgaattcac
      481 gacaggcctt cgaatcaggc accgggcttg cgggtcatgc accaggtgcg cggtccttcg
      541 ggcacctcga cgtcggcggt gacggtgaag ccgagccgct cgtagaaggg gaggttgcgg
      601 ggcgcggagg tctccaggaa ggcgggcacc ccggcgcgct cggccgcctc cactccgggg
      661 agcacgacgg cgctgcccag acccttgccc tggtggtcgg gcgagacgcc gacggtggcc
      721 aggaaccacg cgggctcctt gggccggtgc ggcgccagga ggccttccat ctgttgctgc
      781 gcggccagcc gggaaccgct caactcggcc atgcgcgggc cgatctcggc gaacaccgcc
      841 cccgcttcga cgctctccgg cgtggtccag accgccaccg cggcgccgtc gtccgcgacc
      901 cacaccttgc cgatgtcgag cccgacgcgc gtgaggaaga gttcttgcag ctcggtgacc
      961 cgctcgatgt ggcggtccgg atcgacggtg tggcgcgtgg cggggtagtc ggcgaacgcg
     1021 gcggcgaggg tgcgtacggc cctggggacg tcgtcgcggg tggcgaggcg caccgtgggc
     1081 ttgtactcgg tcatggggcc aggattctcc tcgacgtcac cagcctgctt cagcaggctg
     1141 aagttagtag ctccactgcc gttcagcttg tgccccagtt tgctagggag gtcgcagtat
     1201 ctggccactg ccacctcgtg ctgctcgacg taggtctcgt tgttggcctc cttgattctt
     1261 tccagtctgt agtccacata gtagacgcca ggcatcttga ggttcttagc gggtttcttg
     1321 gatctatatg tggtcttgat gtttgcgatc agatggctcc cgcccacgag cttcagggcc
     1381 atgtcgtttc tgccttccag gccgccgtca gcggggtaca gcgtctcggt gaaggcctcc
     1441 cagccgagtg ttttcttctg catcacaggg ccgttggatg tgaagttcac ccctctgatc
     1501 ttgacgttgt agatgaggca gccgtcctgg aggctggtgt cctgggtagc ggtcagcacg
     1561 cccccgtctt cgtatgtggt gactctctcc catgtgaagc cctcagggaa ggactgcttg
     1621 aagaagtcgg ggatgccctg ggtgtggttg atgaaggtct tgctgccgta gaggaagcta
     1681 gtagccagga tgtcgaaggc gaaggggaga gggccgccct cgaccacctt gattctcatg
     1741 gtctgggtgc cctcgtaggg cttgccttcg ccctcggatg tgcacttgaa gtgatggttg
     1801 tccacggtgc cctccatgta cagcttcatg tgcatgttct ccttaatcag ctcgctcatg
     1861 gtggcctcag gtgcagaggt ttctacaggg aaagagagaa aggctacagt tagtttgctg
     1921 tgatgatgct aagtgccgag gtcagatgtc tatggattgt acacggaatc ccaacgcagg
     1981 aaccaaaaaa cactcaccca aaaaaatggc gctaaggccg agagaaaggc gaagttccgt
     2041 ctacggctat ataccttgaa acaggggcgg cagaagtgac gcaacgcggt tgcgcggacg
     2101 gcttcctgtg ttttcattgg tcgaagcgtt ctgcgcgtgc gcagagattc cccagggaag
     2161 gcgggccctc ttccgaatgt ccaatccgga cattcttccg gttggacctc cgcggtcagc
     2221 gctgtgcccg gcgagagatc acgtggggcg cggaggcggt gctgctgggg cacggccgtc
     2281 cagcctcggc ggccatattt ttgaggggct gttcatctcg ttcacacgct ctgtccgcca
     2341 tgtttgtgag tggaagcgcc attacccctt caagcgactg aaggctgcag ggcctctggt
     2401 ggcccgcatg gggagaccag acccgccagg cccgcctttc cgcactcagt ccgggcttac
     2461 tttattttgt gagacagggt ctcgcctaga ggctccggtg cccgtcagtg ggcagagcgc
     2521 acatcgccca cagtccccga gaagttgggg ggaggggtcg gcaattgaac cggtgcctag
     2581 agaaggtggc gcggggtaaa ctgggaaagt gatgtcgtgt actggctccg cctttttccc
     2641 gagggtgggg gagaaccgta tataagtgca gtagtcgccg tgaacgttct ttttcgcaac
     2701 gggtttgccg ccagaacaca ggtaagtgcc gtgtgtggtt cccgcgggcc tggcctcttt
     2761 acgggttatg gcccttgcgt gccttgaatt acttccacgc ccctggctgc agtacgtgat
     2821 tcttgatccc gagcttcggg ttggaagtgg gtgggagagt tcgaggcctt gcgcttaagg
     2881 agccccttcg cctcgtgctt gagttgaggc ctggcttggg cgctggggcc gccgcgtgcg
     2941 aatctggtgg caccttcgcg cctgtctcgc tgctttcgat aagtctctag ccatttaaaa
     3001 tttttgatga cctgctgcga cgcttttttt ctggcaagat agtcttgtaa atgcgggcca
     3061 agatctgcac actggtattt cggtttttgg ggccgcgggc ggcgacgggg cccgtgcgtc
     3121 ccagcgcaca tgttcggcga ggcggggcct gcgagcgcgg ccaccgagaa tcggacgggg
     3181 gtagtctcaa gctggccggc ctgctctggt gcctggcctc gcgccgccgt gtatcgcccc
     3241 gccctgggcg gcaaggctgg cccggtcggc accagttgcg tgagcggaaa gatggccgct
     3301 tcccggccct gctgcaggga gctcaaaatg gaggacgcgg cgctcgggag agcgggcggg
     3361 tgagtcaccc acacaaagga aaagggcctt tccgtcctca gccgtcgctt catgtgactc
     3421 cacggagtac cgggcgccgt ccaggcacct cgattagttc tcgagctttt ggagtacgtc
     3481 gtctttaggt tggggggagg ggttttatgc gatggagttt ccccatactg agtgggtgga
     3541 gactgaagtt aggccagctt ggcacttgat gtaattctcc ttggaatttg ccctttttga
     3601 gtttggatct tggttcattc tcaagcctca gacagtggtt caaagttttt ttcttccatt
     3661 taaggtgtcg tgaaaactac cccaagctgg cctctggcca ccatggagac agacacactc
     3721 ctgctatggg tactgctgct ctgggttcca ggttccactg gtgacggacg tggcgttcct
     3781 catattgtta tggtggacgc ctacaaacgc tataaatcgg gaggtggttc aggctacacc
     3841 atcgtggagc agtacgagag agccgaggcc agacacagca ccgcgggagg cggtggatcg
     3901 gaacaaaaac tcatctcaga agaggatctg aatgctgtgg gccaggacac gcaggaggtc
     3961 atcgtggtgc cacactcctt gccctttaag gtggtggtga tctcagccat cctggccctg
     4021 gtggtgctca ccatcatctc ccttatcatc ctcatcatgc tttggcagaa gaagccacgt
     4081 taggtcaggc caagcttggg gatcaattct ctagagctcg ctgatcagcc tcgactgtgc
     4141 cttctagttg ccagccatct gttgtttgcc cctcccccgt gccttccttg accctggaag
     4201 gtgccactcc cactgtcctt tcctaataaa atgaggaaat tgcatcgcat tgtctgagta
     4261 ggtgtcattc tattctgggg ggtggggtgg ggcaggacag caagggggag gattgggaag
     4321 acaatagcag gcatgctggg gatgcggtgg gctctatggc ttctgaggcg gaaagaacca
     4381 gctgggccta ctagatagcg gaccccttac cgaaacatcg ccgcattctg cagaggagtc
     4441 gaacttagtg tatgtaaact tctgacccac tggaattgtg atacagtgaa ttataagtga
     4501 aataatctgt ctgtaaacaa ttgttggaaa aatgacttgt gtcatgcaca aagtagatgt
     4561 cctaactgac ttgccaaaac tattgtttgt taacaagaaa tttgtggagt agttgaaaaa
     4621 cgagttttaa tgactccaac ttaagtgtat gtaaacttcc gacttcaact gtatagggat
     4681 ccaattttaa caaaatatta acgcttacaa tttcctgatg cggtattttc tccttacgca
     4741 tctgtgcggt atttcacacc gcatcaggtg gcacttttcg gggaaatgtg cgcggaaccc
     4801 ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct
     4861 gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg
     4921 cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg
     4981 tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc
     5041 tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca
     5101 cttttaaagt tctgctatgt ggcgcggtat tatcccgtat tgacgccggg caagagcaac
     5161 tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa
     5221 agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg
     5281 ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt
     5341 ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg
     5401 aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca acaacgttgc
     5461 gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga
     5521 tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta
     5581 ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc
     5641 cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg
     5701 atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt
     5761 cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa
     5821 ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt
     5881 cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt
     5941 ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt
     6001 tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga
     6061 taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag
     6121 caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata
     6181 agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg
     6241 gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga
     6301 gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca
     6361 ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa
     6421 acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt
     6481 tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac
     6541 ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt
     6601 ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga
     6661 ccgagcgcag cgagtcagtg agcgaggaag cggatcccta tacagttgaa gtcggaagtt
     6721 tacatacacc ttagccaaat acatttaaac tcactttttc acaattcctg acatttaatc
     6781 ctagtaaaaa ttccctgtct tag
//
