LOCUS       sequence-366610-        2692 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 pUC ori)..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2692)
  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 2692)
  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: pVH1002-Tier2(pUC ori)
FEATURES             Location/Qualifiers
     source          1..2692
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(139..727)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(219..238)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(848..1639,1640..1708))
                     /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     1471..1490
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1709..1813)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(1974..1991)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1980..2204
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2350..2366)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2350..2366)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     protein_bind    2522..2543
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2660..2677)
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg
       61 ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa
      121 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg
      181 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc
      241 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc
      301 ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc
      361 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg
      421 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc
      481 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga
      541 gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc
      601 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac
      661 caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg
      721 atctcaagaa gatcctttga tcttttctac ggggattatc aaaaaggatc ttcacctaga
      781 tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt
      841 ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt
      901 catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat
      961 ctggccccag tgctgcaatg ataccgcggc tcccacgctc accggctcca gatttatcag
     1021 caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct
     1081 ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt
     1141 tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg
     1201 cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca
     1261 aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt
     1321 tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat
     1381 gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac
     1441 cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa
     1501 aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt
     1561 tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt
     1621 tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa
     1681 gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt
     1741 atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa
     1801 taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtctaagaa accaagatct
     1861 ccacgcgtgg taccctcgag gagctcttcg aagcggaatt caccatgact agtgctagcg
     1921 gatccaccgg tgtctagaaa gctttgaggc cggcctgcag gcccgctgat cagcctcgac
     1981 tgtgccttct agttgccagc catctgttgt ttgcccctcc cccgtgcctt ccttgaccct
     2041 ggaaggtgcc actcccactg tcctttccta ataaaatgag gaaattgcat cgcattgtct
     2101 gagtaggtgt cattctattc tggggggtgg ggtggggcag gacagcaagg gggaggattg
     2161 ggaagacaat agcaggcatg ctggggatgc ggtgggctct atggtagctt aggaggcgga
     2221 aagaaccagc gcgggagacc gtcagttggt ctccagctat acaggtgcac cgctttataa
     2281 taaaactgat acagagaacg ttatcatgtg tgttggtttt ttttgtgtgt gcggtttaag
     2341 ggagcctcgg tcatagctgt ttcctgtgta aaacgacggc cagcgcggga gacggtcagt
     2401 tcgtctccag ctcagaaatt atgagagcgt ttaataaagt attgactatg gttgacaaaa
     2461 cgtgtgttgg agttttttgt gtgctttgtc atcaactgag ccagtaggtc gagggtgcct
     2521 aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa
     2581 acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta
     2641 ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cg
//
