LOCUS       sequence-366220-        4981 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 YPet marker and BlastR 
            resistance gene 
            (5'ITR-A1-pA::A2-pA::PRPBSA-YPet-p2A-BlastR-pA-3'ITR).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4981)
  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 4981)
  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: pTS2343_Tier3(SB)-YPet_Blast
FEATURES             Location/Qualifiers
     source          1..4981
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(190..207)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    196..420
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(566..582)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(566..582)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1575..1594)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     CDS             2013..2069
                     /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"
     protein_bind    2611..2632
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(3034..3622)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3114..3133)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3743..4534,4535..4603))
                     /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     4366..4385
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4604..4708)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 acatacacta agttcgactc ctctgcagaa tgcggcgatg tttcggtaag gggtccgcta
       61 tctagtctta agagatctac gcgtggtacc ctcgaggagc tcttcgaagc ggaattcacc
      121 atgactagtg ctagcggatc caccggtgtc tagaaagctt tgaggccggc ctgcaggccc
      181 gctgatcagc ctcgactgtg ccttctagtt gccagccatc tgttgtttgc ccctcccccg
      241 tgccttcctt gaccctggaa ggtgccactc ccactgtcct ttcctaataa aatgaggaaa
      301 ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg gggtggggtg gggcaggaca
      361 gcaaggggga ggattgggaa gacaatagca ggcatgctgg ggatgcggtg ggctctatgg
      421 tagcttagga ggcggaaaga accagcgcgg gagaccgtca gttggtctcc agctatacag
      481 gtgcaccgct ttataataaa actgatacag agaacgttat catgtgtgtt ggtttttttt
      541 gtgtgtgcgg tttaagggag cctcggtcat agctgtttcc tgtgtaaaac gacggccagc
      601 gcgtggtacc ctcgagcgtg accctgtctc acaaaataaa gtaagcccgg actgagtgcg
      661 gaaaggcggg cctggcgggt ctggactccc catgcgggcc accagaggcc ctgcagcctt
      721 cagtcgcttg aaggggtaat ggcgcttcca ctcacaaaca tggcggacag agcgtgtgaa
      781 cgagatgaac agcccctcaa aaatatggcc gccgaggctg gacggccgtg ccccagcagc
      841 accgcctccg cgccccacgt gatctctcgc cgggcacagc gctgaccgcg gaggtccaac
      901 cggaagaatg tccggattgg acattcggaa gagggcccgc cttccctggg gaatctctgc
      961 gcacgcgcag aacgcttcga ccaatgaaaa cacaggaagc cgtccgcgca accgcgttgc
     1021 gtcacttctg ccgcccctgt ttcaaggtat atagccgtag acggaacttc gcctttctct
     1081 cggccttagc gccatttttt tgggtgagtg ttttttggtt cctgcgttgg gattccgtgt
     1141 acaatccata gacatctgac ctcggcactt agcatcatca cagcaaacta actgtagcct
     1201 ttctctcttt ccctgtagaa acctctgcac ctagggagct cgaattcacc atgactagtg
     1261 gtggttctgg tgtgagcaaa ggcgaagagc tgttcaccgg cgtggtgccc atcctggtgg
     1321 agctggacgg cgacgtgaac ggccacaagt tcagcgtgag cggcgagggc gagggcgacg
     1381 ccacctacgg caagctgacc ctgaagctgc tgtgcaccac cggcaagctg cccgtgccct
     1441 ggcccaccct ggtgaccacc ctgggctacg gcgtgcagtg cttcgcccgg taccccgacc
     1501 acatgaagca gcacgacttc ttcaagagcg ccatgcccga gggctacgtg caggagcgga
     1561 ccatcttctt caaggacgac ggcaactaca agacccgggc cgaggtgaag ttcgagggcg
     1621 acaccctggt gaaccggatc gagctgaagg gcatcgactt caaggaggac ggcaacatcc
     1681 tgggccacaa gctggagtac aactacaaca gccacaacgt gtacatcacc gccgacaagc
     1741 agaagaacgg catcaaggcc aacttcaaga tccggcacaa catcgaggac ggcggcgtgc
     1801 agctggccga ccactaccag cagaacaccc ccatcggcga cggccccgtg ctgctgcccg
     1861 acaaccacta cctgagctac cagagcgccc tgttcaagga ccccaacgag aagcgggacc
     1921 acatggtgct gctggagttc ctgaccgccg ccggcatcac cgagggcatg gacgagctgt
     1981 acaaggctag tggtggttct ggtggaagcg gagctactaa cttcagcctg ctgaagcagg
     2041 ctggtgacgt ggaggagaac cctggacctg ctagtggtgg ttctggtttg tctcaagaag
     2101 aatccaccct cattgaaaga gcaacggcta caatcaacag catccccatc tctgaagact
     2161 acagcgtcgc cagcgcagct ctctctagcg acggccgcat cttcactggt gtcaatgtat
     2221 atcattttac tgggggacct tgtgcagaac tcgtggtgct gggcactgct gctgctgcgg
     2281 cagctggcaa cctgacttgt atcgtcgcga tcggaaatga gaacaggggc atcttgagcc
     2341 cctgcggacg gtgccgacag gtgcttctcg atctgcatcc tgggatcaaa gccatagtga
     2401 aggacagtga tggacagccg acggcagttg ggattcgtga attgctgccc tctggttatg
     2461 tgtgggaggg cgctagcgga tccaccggtg tctagaaagc tcagaaatta tgagagcgtt
     2521 taataaagta ttgactatgg ttgacaaaac gtgtgttgga gttttttgtg tgctttgtca
     2581 tcaactgagc cagtaggtcg agggtgccta atgagtgagc taactcacat taattgcgtt
     2641 gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgctag ctactcggga
     2701 ccccttaccg aaacatcgcc gcattctgca gaggagtcga gtgtatgtaa acttctgacc
     2761 cactgggaat gtgatgaaag aaataaaagc tgaaatgaat cattctctct actattattc
     2821 tgatatttca cattcttaaa ataaagtggt gatcctaact gacctaagac agggaatttt
     2881 tactaggatt aaatgtcagg aattgtgaaa aagtgagttt aaatgtattt ggctaaggtg
     2941 tatgtaaact tccgacttca actgtaccat ggacatgtga gcaaaaggcc agcaaaaggc
     3001 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     3061 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     3121 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     3181 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     3241 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     3301 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     3361 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     3421 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt
     3481 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     3541 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     3601 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacgggga ttatcaaaaa
     3661 ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat
     3721 atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga
     3781 tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac
     3841 gggagggctt accatctggc cccagtgctg caatgatacc gcggctccca cgctcaccgg
     3901 ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg
     3961 caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt
     4021 cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct
     4081 cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat
     4141 cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta
     4201 agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca
     4261 tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat
     4321 agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac
     4381 atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa
     4441 ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt
     4501 cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg
     4561 caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat
     4621 attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt
     4681 agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtct
     4741 aagaaaccaa gatcagcggc cgctacagtt gaagtcggaa gtttacatac agttaagttg
     4801 gagtcattaa aactcgtttt tcaactactc cacaaatttc ttgttaacaa acaatagttt
     4861 tggcaagtca gttaggacat ctactttgtg catgacacaa gtcatttttc caacaattgt
     4921 ttacagacag attatttcac ttataattca ctgtatcaca attccagtgg gtcagaagtt
     4981 t
//
