LOCUS       sequence-365621-        3990 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Tier-2 vector encoding PhCMV-driven secreted Nluc 
            (A1-pA::PhCMV-IgkSS-Nluc-pA::A3-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: pTS1032
FEATURES             Location/Qualifiers
     source          1..3990
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(30..47)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(201..789)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(281..300)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(910..1701,1702..1770))
                     /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     1533..1552
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1771..1875)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(2036..2053)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2042..2266
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     enhancer        2311..2690
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2691..2894
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2845..2865
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2939..2958
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2939..2957
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     sig_peptide     2998..3054
                     /codon_start=1
                     /product="leader sequence from mouse immunoglobulin kappa 
                     light chain"
                     /label=Ig-kappa leader
                     /translation="ETDTLLLWVLLLWVPGSTG"
     CDS             3076..3588
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="MVFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQR
                     IVLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGV
                     TPNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGW
                     RLCERILA"
     primer_bind     complement(3710..3726)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3710..3726)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     protein_bind    3882..3903
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
ORIGIN
        1 ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt
       61 cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca
      121 ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa
      181 aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat
      241 cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc
      301 cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc
      361 gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt
      421 tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac
      481 cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg
      541 ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca
      601 gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc
      661 gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa
      721 accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa
      781 ggatctcaag aagatccttt gatcttttct acggggatta tcaaaaagga tcttcaccta
      841 gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg
      901 gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg
      961 ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc
     1021 atctggcccc agtgctgcaa tgataccgcg gctcccacgc tcaccggctc cagatttatc
     1081 agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc
     1141 ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag
     1201 tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat
     1261 ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg
     1321 caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt
     1381 gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag
     1441 atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg
     1501 accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt
     1561 aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct
     1621 gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac
     1681 tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat
     1741 aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat
     1801 ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca
     1861 aataggggtt ccgcgcacat ttccccgaaa agtgccacct gacgtctaag aaaccaagat
     1921 ctccacgcgt ggtaccctcg aggagctctt cgaagcggaa ttcaccatga ctagtgctag
     1981 cggatccacc ggtgtctaga aagctttgag gccggcctgc aggcccgctg atcagcctcg
     2041 actgtgcctt ctagttgcca gccatctgtt gtttgcccct cccccgtgcc ttccttgacc
     2101 ctggaaggtg ccactcccac tgtcctttcc taataaaatg aggaaattgc atcgcattgt
     2161 ctgagtaggt gtcattctat tctggggggt ggggtggggc aggacagcaa gggggaggat
     2221 tgggaagaca atagcaggca tgctggggat gcggtgggct ctatggtagc ttaggaggcg
     2281 gaaagaacca gcgcgtggta ccctcgagtt gacattgatt attgagtagt tattaatagt
     2341 aatcaattac ggggtcatta gttcatagcc catatatgga gttccgcgtt acataactta
     2401 cggtaaatgg cccgcctggc tgaccgccca acgacccccg cccattgacg tcaataatga
     2461 cgtatgttcc catagtaacg ccaataggga ctttccattg acgtcaatgg gtggagtatt
     2521 tacggtaaac tgcccacttg gcagtacatc aagtgtatca tatgccaagt acgcccccta
     2581 ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc ccagtacatg accttatggg
     2641 actttcctac ttggcagtac atctacgtat tagtcatcgc tattaccatg gtgatgcggt
     2701 tttggcagta catcaatggg cgtggatagc ggtttgactc acggggattt ccaagtctcc
     2761 accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat
     2821 gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg tgggaggtct
     2881 atataagcag agctctctgg ctaactagag aacccactgc ttactggctt atcgaaatta
     2941 atacgactca ctatagggag acccaagctg ttcgaagcgg aattcaccat gactagtgag
     3001 acagacacac tcctgctatg ggtactgctg ctctgggttc caggttccac tggtgacgct
     3061 agtggtggtt ctggtatggt cttcacactc gaagatttcg ttggggactg gcgacagaca
     3121 gccggctaca acctggacca agtccttgaa cagggaggtg tgtccagttt gtttcagaat
     3181 ctcggggtgt ccgtaactcc gatccaaagg attgtcctga gcggtgaaaa tgggctgaag
     3241 atcgacatcc atgtcatcat cccgtatgaa ggtctgagcg gcgaccaaat gggccagatc
     3301 gaaaaaattt ttaaggtggt gtaccctgtg gatgatcatc actttaaggt gatcctgcac
     3361 tatggcacac tggtaatcga cggggttacg ccgaacatga tcgactattt cggacggccg
     3421 tatgaaggca tcgccgtgtt cgacggcaaa aagatcactg taacagggac cctgtggaac
     3481 ggcaacaaaa ttatcgacga gcgcctgatc aaccccgacg gctccctgct gttccgagta
     3541 accatcaacg gagtgaccgg ctggcggctg tgcgaacgca ttctggcggc tagcggatcc
     3601 accggtgtct agaaagctat acaggtgcac cgctttataa taaaactgat acagagaacg
     3661 ttatcatgtg tgttggtttt ttttgtgtgt gcggtttaag ggagcctcgg tcatagctgt
     3721 ttcctgtgta aaacgacggc cagcgcggga gacggtcagt tcgtctccag ctcagaaatt
     3781 atgagagcgt ttaataaagt attgactatg gttgacaaaa cgtgtgttgg agttttttgt
     3841 gtgctttgtc atcaactgag ccagtaggtc gagggtgcct aatgagtgag ctaactcaca
     3901 ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat
     3961 taatgaatcg gccaacgcgc ggggagaggc
//
