LOCUS       sequence-366789-        4375 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Tier-3 vector for stable stable integration of up to three cassettes
            via sleeping beauty transposase including a PuroR resistance gene 
            (5'ITR-A1-pA::A2-pA::PRPBSA-PuroR-pA-3'ITR).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4375)
  AUTHORS   Haellman V, Strittmatter T, Bertschi A, Stucheli P, Fussenegger M
  TITLE     A versatile plasmid architecture for mammalian synthetic biology 
            (VAMSyB).
  JOURNAL   Metab Eng. 2021 Jul;66:41-50. doi: 10.1016/j.ymben.2021.04.003. Epub
            2021 Apr 18.
  PUBMED    33857582
REFERENCE   2  (bases 1 to 4375)
  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: "Metab 
            Eng."; date: "2021-07"; volume: "66:41-50. doi"; pages: " 
            10.1016/j.ymben.2021.04.003. Epub 2021 Apr 18"
COMMENT     Sequence Label: pTS2341_Tier3(SB)-Puro
FEATURES             Location/Qualifiers
     source          1..4375
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(593..1384,1385..1453))
                     /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     1216..1235
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1454..1558)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(2021..2038)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2027..2251
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2397..2413)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2397..2413)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             3103..3696
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="TEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIERV
                     TELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLAA
                     QQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETSA
                     PRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     3596..3616
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     protein_bind    3836..3857
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(join(4259..4375,1..472))
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4339..4358)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg
       61 aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg
      121 ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg
      181 taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac
      241 tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg
      301 gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt
      361 taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg
      421 tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc
      481 tttgatcttt tctacgggga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
      541 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat
      601 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct
      661 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg
      721 caatgatacc gcggctccca cgctcaccgg ctccagattt atcagcaata aaccagccag
      781 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta
      841 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg
      901 ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg
      961 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct
     1021 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta
     1081 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg
     1141 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc
     1201 cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg
     1261 gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga
     1321 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg
     1381 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat
     1441 gttgaatact catactcttc ctttttcaat attattgaag catttatcag ggttattgtc
     1501 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca
     1561 catttccccg aaaagtgcca cctgacgtct aagaaaccaa gatcagcggc cgctacagtt
     1621 gaagtcggaa gtttacatac agttaagttg gagtcattaa aactcgtttt tcaactactc
     1681 cacaaatttc ttgttaacaa acaatagttt tggcaagtca gttaggacat ctactttgtg
     1741 catgacacaa gtcatttttc caacaattgt ttacagacag attatttcac ttataattca
     1801 ctgtatcaca attccagtgg gtcagaagtt tacatacact aagttcgact cctctgcaga
     1861 atgcggcgat gtttcggtaa ggggtccgct atctagtctt aagagatcta cgcgtggtac
     1921 cctcgaggag ctcttcgaag cggaattcac catgactagt gctagcggat ccaccggtgt
     1981 ctagaaagct ttgaggccgg cctgcaggcc cgctgatcag cctcgactgt gccttctagt
     2041 tgccagccat ctgttgtttg cccctccccc gtgccttcct tgaccctgga aggtgccact
     2101 cccactgtcc tttcctaata aaatgaggaa attgcatcgc attgtctgag taggtgtcat
     2161 tctattctgg ggggtggggt ggggcaggac agcaaggggg aggattggga agacaatagc
     2221 aggcatgctg gggatgcggt gggctctatg gtagcttagg aggcggaaag aaccagcgcg
     2281 ggagaccgtc agttggtctc cagctataca ggtgcaccgc tttataataa aactgataca
     2341 gagaacgtta tcatgtgtgt tggttttttt tgtgtgtgcg gtttaaggga gcctcggtca
     2401 tagctgtttc ctgtgtaaaa cgacggccag cgcgtggtac cctcgagcgt gaccctgtct
     2461 cacaaaataa agtaagcccg gactgagtgc ggaaaggcgg gcctggcggg tctggactcc
     2521 ccatgcgggc caccagaggc cctgcagcct tcagtcgctt gaaggggtaa tggcgcttcc
     2581 actcacaaac atggcggaca gagcgtgtga acgagatgaa cagcccctca aaaatatggc
     2641 cgccgaggct ggacggccgt gccccagcag caccgcctcc gcgccccacg tgatctctcg
     2701 ccgggcacag cgctgaccgc ggaggtccaa ccggaagaat gtccggattg gacattcgga
     2761 agagggcccg ccttccctgg ggaatctctg cgcacgcgca gaacgcttcg accaatgaaa
     2821 acacaggaag ccgtccgcgc aaccgcgttg cgtcacttct gccgcccctg tttcaaggta
     2881 tatagccgta gacggaactt cgcctttctc tcggccttag cgccattttt ttgggtgagt
     2941 gttttttggt tcctgcgttg ggattccgtg tacaatccat agacatctga cctcggcact
     3001 tagcatcatc acagcaaact aactgtagcc tttctctctt tccctgtaga aacctctgca
     3061 cctagggagc tcgaattcac catgactagt ggtggttctg gtaccgagta caagcccacg
     3121 gtgcgcctcg ccacccgcga cgacgtcccc cgggccgtac gcaccctcgc cgccgcgttc
     3181 gccgactacc ccgccacgcg ccacaccgtc gacccggacc gccacatcga gcgggtcacc
     3241 gagctgcaag aactcttcct cacgcgcgtc gggctcgaca tcggcaaggt gtgggtcgcg
     3301 gacgacggcg ccgcggtggc ggtctggacc acgccggaga gcgtcgaagc gggggcggtg
     3361 ttcgccgaga tcggcccgcg catggccgag ttgagcggtt cccggctggc cgcgcagcaa
     3421 cagatggaag gcctcctggc gccgcaccgg cccaaggagc ccgcgtggtt cctggccacc
     3481 gtcggcgtgt cgcccgacca ccagggcaag ggtctgggca gcgccgtcgt gctccccgga
     3541 gtggaggcgg ccgagcgcgc cggggtgccc gccttcctgg agacatccgc gccccgcaac
     3601 ctccccttct acgagcggct cggcttcacc gtcaccgccg acgtcgaggt gcccgaagga
     3661 ccgcgcacct ggtgcatgac ccgcaagccc ggtgccgcta gcggatccac cggtgtctag
     3721 aaagctcaga aattatgaga gcgtttaata aagtattgac tatggttgac aaaacgtgtg
     3781 ttggagtttt ttgtgtgctt tgtcatcaac tgagccagta ggtcgagggt gcctaatgag
     3841 tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt
     3901 cgtgccagct gctagctact cgggacccct taccgaaaca tcgccgcatt ctgcagagga
     3961 gtcgagtgta tgtaaacttc tgacccactg ggaatgtgat gaaagaaata aaagctgaaa
     4021 tgaatcattc tctctactat tattctgata tttcacattc ttaaaataaa gtggtgatcc
     4081 taactgacct aagacaggga atttttacta ggattaaatg tcaggaattg tgaaaaagtg
     4141 agtttaaatg tatttggcta aggtgtatgt aaacttccga cttcaactgt accatggaca
     4201 tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt
     4261 tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc
     4321 gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc ctcgt
//
