LOCUS       sequence-366207-        2742 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Tier-2 expression vector (A1-pA::A2-pA::A3-pA). The tier-2 
            expression vector consists of a custom MCS cassette containing the 
            three Tier-1 compatible acceptor cassettes (A1, A2, and A3) each 
            with its own non-homologous polyadenylation (pA) signals in a 
            minimal backbone (AmpR and ColE1 ori). .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2742)
  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 2742)
  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: pVH1001-Tier2(ColE1 ori)
FEATURES             Location/Qualifiers
     source          1..2742
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(25..816,817..885))
                     /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     648..667
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(886..990)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(1151..1168)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1157..1381
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(1527..1543)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1527..1543)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     protein_bind    1699..1720
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1837..1854)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2008..2596)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2088..2107)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc
       61 gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat
      121 acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcggctcc cacgctcacc
      181 ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc
      241 tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag
      301 ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg
      361 ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg
      421 atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag
      481 taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt
      541 catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga
      601 atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc
      661 acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc
      721 aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc
      781 ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc
      841 cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca
      901 atattattga agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat
      961 ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgt
     1021 ctaagaaacc aagatctcca cgcgtggtac cctcgaggag ctcttcgaag cggaattcac
     1081 catgactagt gctagcggat ccaccggtgt ctagaaagct ttgaggccgg cctgcaggcc
     1141 cgctgatcag cctcgactgt gccttctagt tgccagccat ctgttgtttg cccctccccc
     1201 gtgccttcct tgaccctgga aggtgccact cccactgtcc tttcctaata aaatgaggaa
     1261 attgcatcgc attgtctgag taggtgtcat tctattctgg ggggtggggt ggggcaggac
     1321 agcaaggggg aggattggga agacaatagc aggcatgctg gggatgcggt gggctctatg
     1381 gtagcttagg aggcggaaag aaccagcgcg ggagaccgtc agttggtctc cagctataca
     1441 ggtgcaccgc tttataataa aactgataca gagaacgtta tcatgtgtgt tggttttttt
     1501 tgtgtgtgcg gtttaaggga gcctcggtca tagctgtttc ctgtgtaaaa cgacggccag
     1561 cgcgggagac ggtcagttcg tctccagctc agaaattatg agagcgttta ataaagtatt
     1621 gactatggtt gacaaaacgt gtgttggagt tttttgtgtg ctttgtcatc aactgagcca
     1681 gtaggtcgag ggtgcctaat gagtgagcta actcacatta attgcgttgc gctcactgcc
     1741 cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg
     1801 gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc
     1861 ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac
     1921 agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa
     1981 ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca
     2041 caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc
     2101 gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata
     2161 cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta
     2221 tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca
     2281 gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga
     2341 cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg
     2401 tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg
     2461 tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg
     2521 caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag
     2581 aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa
     2641 cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat
     2701 ccttttaaat taaaaatgaa gttttaaatc aatctaaagt at
//
