LOCUS       sequence-479773-        5543 bp    DNA     circular SYN 05-AUG-2025
DEFINITION  Express NanoLuc-CISH Fusion Protein in Mammalian Cells under a CMV 
            promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5543)
  AUTHORS   Promega
  TITLE     Promega Plasmid Collection
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 5543)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Aug 5, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: NanoLuc-CISH Fusion Vector
FEATURES             Location/Qualifiers
     source          1..5543
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        138..517
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        518..721
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     672..692
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     intron          857..989
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     972..991
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     primer_bind     1033..1052
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1033..1051
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1065..1577
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="MVFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQR
                     IVLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGV
                     TPNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGW
                     RLCERILA"
     primer_bind     complement(2486..2504)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     polyA_signal    2541..2662
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2552..2571)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2606..2625
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        2907..3264
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2907..3098
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2930..2950)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3115..3250
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     promoter        3279..3326
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3358..4152
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3412..3431)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4022..4041
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4216..4264
                     /note="synthetic polyadenylation signal"
     rep_origin      complement(4503..5091)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4583..4602)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     misc_feature    5207..5490
                     /label=cer region
                     /note="ColE1-derived recombination site that helps to 
                     maintain plasmids as monomers"
ORIGIN
        1 tcaatattgg ccattagcca tattattcat tggttatata gcataaatca atattggcta
       61 ttggccattg catacgttgt atctatatca taatatgtac atttatattg gctcatgtcc
      121 aatatgaccg ccatgttggc attgattatt gactagttat taatagtaat caattacggg
      181 gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc
      241 gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat
      301 agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc
      361 ccacttggca gtacatcaag tgtatcatat gccaagtccg ccccctattg acgtcaatga
      421 cggtaaatgg cccgcctggc attatgccca gtacatgacc ttacgggact ttcctacttg
      481 gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt ggcagtacac
      541 caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt
      601 caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaataaccc
      661 cgccccgttg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata taagcagagc
      721 tggtttagtg aaccgtcaga tcactagaag ctttattgcg gtagtttatc acagttaaat
      781 tgctaacgca gtcagtgctt ctgacacaac agtctcgaac ttaagctgca gaagttggtc
      841 gtgaggcact gggcaggtaa gtatcaaggt tacaagacag gtttaaggag accaatagaa
      901 actgggcttg tcgagacaga gaagactctt gcgtttctga taggcaccta ttggtcttac
      961 tgacatccac tttgcctttc tctccacagg tgtccactcc cagttcaatt acagctctta
     1021 aggctagagt attaatacga ctcactatag ggctagcgct caccatggtc ttcacactcg
     1081 aagatttcgt tggggactgg cgacagacag ccggctacaa cctggaccaa gtccttgaac
     1141 agggaggtgt gtccagtttg tttcagaatc tcggggtgtc cgtaactccg atccaaagga
     1201 ttgtcctgag cggtgaaaat gggctgaaga tcgacatcca tgtcatcatc ccgtatgaag
     1261 gtctgagcgg cgaccaaatg ggccagatcg aaaaaatttt taaggtggtg taccctgtgg
     1321 atgatcatca ctttaaggtg atcctgcact atggcacact ggtaatcgac ggggttacgc
     1381 cgaacatgat cgactatttc ggacggccgt atgaaggcat cgccgtgttc gacggcaaaa
     1441 agatcactgt aacagggacc ctgtggaacg gcaacaaaat tatcgacgag cgcctgatca
     1501 accccgacgg ctccctgctg ttccgagtaa ccatcaacgg agtgaccggc tggcggctgt
     1561 gcgaacgcat tctggcgggc tcgagcggcg cgatcgccat ggttctctgc gttcagggac
     1621 ctcgtccttt gctggctgtt gagcggactg gtcagcggcc actgtgggct ccgtccctgg
     1681 aactgcccaa gccagtcatg cagccattgc ctgctggtgc cttcctcgaa gaggttgcag
     1741 agggtacccc agcccagaca gagagtgagc caaaggtgct ggacccagag gaggatctgc
     1801 tgtgcatagc caagaccttc tcctaccttc gggaatctgg ctggtattgg ggttccatta
     1861 cggccagcga ggcccgacaa cacctgcaga agatgccaga aggcacgttc ttagtacgtg
     1921 acagcacgca ccccagctac ctgttcacgc tgtcagtgaa aaccactcgt ggcccaacca
     1981 atgtacgcat tgagtatgcc gactccagct tccgtctgga ctccaactgc ttgtccaggc
     2041 cacgcatcct ggcctttccg gatgtggtca gccttgtgca gcactatgtg gcctcctgca
     2101 ctgctgatac ccgaagcgac agcccagatc ctgctcccac cccagccctg cctatgccta
     2161 aggaggatgc gcctagtgac ccagcactgc ctgctcctcc accagctact gctgtacacc
     2221 taaaactggt gcagcccttt gtacgcagaa gcagtgcacg cagcctgcaa cacctgtgcc
     2281 gccttgttat caaccgtctg gtggccgatg ttgactgcct gccactgccc cggcgcatgg
     2341 ctgactacct ccgacagtac cccttccagc tcgtttaaac gaattcgggc tcggtacccg
     2401 gggatcctct agagtcgacc tgcaggcatg caagctgatc cggctgctaa caaagcccga
     2461 aaggaagctg agttggctgc tgccaccgct gagcaataac tagcataacc ccttggggcg
     2521 gccgcttcga gcagacatga taagatacat tgatgagttt ggacaaacca caactagaat
     2581 gcagtgaaaa aaatgcttta tttgtgaaat ttgtgatgct attgctttat ttgtaaccat
     2641 tataagctgc aataaacaag ttaacaacaa caattgcatt cattttatgt ttcaggttca
     2701 gggggagatg tgggaggttt ttttaagcaa gtaaaacctc tacaaatgtg gtaaaatcga
     2761 attttaacaa aatattaacg cttacaattt cctgatgcgg tattttctcc ttacgcatct
     2821 gtgcggtatt tcacaccgca tacgcggatc tgcgcagcac catggcctga aataacctct
     2881 gaaagaggaa cttggttagg taccttctga ggcggaaaga accagctgtg gaatgtgtgt
     2941 cagttagggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat
     3001 ctcaattagt cagcaaccag gtgtggaaag tccccaggct ccccagcagg cagaagtatg
     3061 caaagcatgc atctcaatta gtcagcaacc atagtcccgc ccctaactcc gcccatcccg
     3121 cccctaactc cgcccagttc cgcccattct ccgccccatg gctgactaat tttttttatt
     3181 tatgcagagg ccgaggccgc ctcggcctct gagctattcc agaagtagtg aggaggcttt
     3241 tttggaggcc taggcttttg caaaaagctt aattaactgt tgacaattaa tcatcggcat
     3301 agtatatcgg catagtataa tacgacaagg tgaggaacta aacccaggag gcagatcatg
     3361 attgaacaag atggattgca cgcaggttct ccggccgctt gggtggagag gctattcggc
     3421 tatgactggg cacaacagac aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg
     3481 caggggcgcc cggttctttt tgtcaagacc gacctgtccg gtgccctgaa tgaactgcag
     3541 gacgaggcag cgcggctatc gtggctggcc acgacgggcg ttccttgcgc agctgtgctc
     3601 gacgttgtca ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc ggggcaggat
     3661 ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca tcatggctga tgcaatgcgg
     3721 cggctgcata cgcttgatcc ggctacctgc ccattcgacc accaagcgaa acatcgcatc
     3781 gagcgagcac gtactcggat ggaagccggt cttgtcgatc aggatgatct ggacgaagag
     3841 catcaggggc tcgcgccagc cgaactgttc gccaggctca aggcgcgcat gcccgacggc
     3901 gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga atatcatggt ggaaaatggc
     3961 cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata
     4021 gcgttggcta cccgtgatat tgctgaagag cttggcggcg aatgggctga ccgcttcctc
     4081 gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg ccttcttgac
     4141 gagttcttct gagcgggact ctggggttcg aaatgaccga ccaagcgacg cccaacctgc
     4201 catcacgatg gccgcaataa aatatcttta ttttcattac atctgtgtgt tggttttttg
     4261 tgtgaatcga tagcgataag gatcctcttt gcgcttgcgt tttcccttgt ccagatagcc
     4321 cagtagctga cattcatccg gggtcagcac cgtttctgcg gactggcttt ctacccggta
     4381 tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag
     4441 aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg
     4501 tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg
     4561 tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg
     4621 cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga
     4681 agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc
     4741 tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt
     4801 aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact
     4861 ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg
     4921 cctaactacg gctacactag aaggacagta tttggtatct gcgctctgct gaagccagtt
     4981 accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt
     5041 ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatttca agaagatcct
     5101 ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg
     5161 gtcatgagat tatcaaaaag gatcttcacc tagatccttt tatagtccgg aaatacagga
     5221 acgcacgctg gatggccctt cgctgggatg gtgaaaccat gaaaaatggc agcttcagtg
     5281 gattaagtgg gggtaatgtg gcctgtaccc tctggttgca taggtattca tacggttaaa
     5341 atttatcagg cgcgattgcg gcagtttttc gggtggtttg ttgccatttt tacctgtctg
     5401 ctgccgtgat cgcgctgaac gcgttttagc ggtgcgtaca attaagggat tatggtaaat
     5461 ccacttactg tctgccctcg tagccatcga gataaaccgc agtactccgg ccacgatgcg
     5521 tccggcgtag aggatcgaga tct
//
