LOCUS       sequence-365610-        3990 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Tier-2 vector encoding PhCMV-driven secreted Nluc 
            (A1-pA::A2-pA::PhCMV-IgkSS-Nluc-pA)..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3990)
  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 3990)
  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: pTS1033
FEATURES             Location/Qualifiers
     source          1..3990
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(36..827,828..896))
                     /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     659..678
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(897..1001)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(1162..1179)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1168..1392
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(1538..1554)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1538..1554)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     enhancer        1591..1970
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1971..2174
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2125..2145
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2219..2238
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2219..2237
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     sig_peptide     2278..2334
                     /codon_start=1
                     /product="leader sequence from mouse immunoglobulin kappa 
                     light chain"
                     /label=Ig-kappa leader
                     /translation="ETDTLLLWVLLLWVPGSTG"
     CDS             2356..2868
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="MVFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQR
                     IVLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGV
                     TPNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGW
                     RLCERILA"
     protein_bind    3008..3029
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3146..3163)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3317..3905)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3397..3416)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca
       61 cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag
      121 ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcggctc
      181 ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc
      241 agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct
      301 agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc
      361 gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg
      421 cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc
      481 gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat
      541 tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag
      601 tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat
      661 aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg
      721 cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca
      781 cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga
      841 aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc
      901 ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag cggatacata
      961 tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg
     1021 ccacctgacg tctaagaaac caagatctcc acgcgtggta ccctcgagga gctcttcgaa
     1081 gcggaattca ccatgactag tgctagcgga tccaccggtg tctagaaagc tttgaggccg
     1141 gcctgcaggc ccgctgatca gcctcgactg tgccttctag ttgccagcca tctgttgttt
     1201 gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc ctttcctaat
     1261 aaaatgagga aattgcatcg cattgtctga gtaggtgtca ttctattctg gggggtgggg
     1321 tggggcagga cagcaagggg gaggattggg aagacaatag caggcatgct ggggatgcgg
     1381 tgggctctat ggtagcttag gaggcggaaa gaaccagcgc gggagaccgt cagttggtct
     1441 ccagctatac aggtgcaccg ctttataata aaactgatac agagaacgtt atcatgtgtg
     1501 ttggtttttt ttgtgtgtgc ggtttaaggg agcctcggtc atagctgttt cctgtgtaaa
     1561 acgacggcca gcgcgtggta ccctcgagtt gacattgatt attgagtagt tattaatagt
     1621 aatcaattac ggggtcatta gttcatagcc catatatgga gttccgcgtt acataactta
     1681 cggtaaatgg cccgcctggc tgaccgccca acgacccccg cccattgacg tcaataatga
     1741 cgtatgttcc catagtaacg ccaataggga ctttccattg acgtcaatgg gtggagtatt
     1801 tacggtaaac tgcccacttg gcagtacatc aagtgtatca tatgccaagt acgcccccta
     1861 ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc ccagtacatg accttatggg
     1921 actttcctac ttggcagtac atctacgtat tagtcatcgc tattaccatg gtgatgcggt
     1981 tttggcagta catcaatggg cgtggatagc ggtttgactc acggggattt ccaagtctcc
     2041 accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat
     2101 gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg tgggaggtct
     2161 atataagcag agctctctgg ctaactagag aacccactgc ttactggctt atcgaaatta
     2221 atacgactca ctatagggag acccaagctg ttcgaagcgg aattcaccat gactagtgag
     2281 acagacacac tcctgctatg ggtactgctg ctctgggttc caggttccac tggtgacgct
     2341 agtggtggtt ctggtatggt cttcacactc gaagatttcg ttggggactg gcgacagaca
     2401 gccggctaca acctggacca agtccttgaa cagggaggtg tgtccagttt gtttcagaat
     2461 ctcggggtgt ccgtaactcc gatccaaagg attgtcctga gcggtgaaaa tgggctgaag
     2521 atcgacatcc atgtcatcat cccgtatgaa ggtctgagcg gcgaccaaat gggccagatc
     2581 gaaaaaattt ttaaggtggt gtaccctgtg gatgatcatc actttaaggt gatcctgcac
     2641 tatggcacac tggtaatcga cggggttacg ccgaacatga tcgactattt cggacggccg
     2701 tatgaaggca tcgccgtgtt cgacggcaaa aagatcactg taacagggac cctgtggaac
     2761 ggcaacaaaa ttatcgacga gcgcctgatc aaccccgacg gctccctgct gttccgagta
     2821 accatcaacg gagtgaccgg ctggcggctg tgcgaacgca ttctggcggc tagcggatcc
     2881 accggtgtct agaaagctca gaaattatga gagcgtttaa taaagtattg actatggttg
     2941 acaaaacgtg tgttggagtt ttttgtgtgc tttgtcatca actgagccag taggtcgagg
     3001 gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt
     3061 cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt
     3121 tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc
     3181 tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg
     3241 ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg
     3301 ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac
     3361 gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg
     3421 gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct
     3481 ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg
     3541 tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct
     3601 gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac
     3661 tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt
     3721 tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc
     3781 tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca
     3841 ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat
     3901 ctcaagaaga tcctttgatc ttttctacgg ggattatcaa aaaggatctt cacctagatc
     3961 cttttaaatt aaaaatgaag ttttaaatca
//
