LOCUS       sequence-471837-        6617 bp    DNA     circular SYN 18-JUN-2025
DEFINITION  Express IRES in Mammalian Cells under a CMV promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6617)
  TITLE     Promega Early Access Collection
REFERENCE   2  (bases 1 to 6617)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jun 18, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     Sequence Label: pBiT4.1-C [HiBiT-IRES-luc2/CMV/Blast] Vector
FEATURES             Location/Qualifiers
     source          1..6617
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    24..105
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     enhancer        338..641
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        642..845
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     796..816
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     intron          981..1113
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     1096..1115
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     primer_bind     1157..1176
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1157..1175
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_feature    1365..1937
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     misc_feature    1388..1939
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(1532..1549)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     1759..1778
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             1939..3591
                     /codon_start=1
                     /product="firefly luciferase"
                     /label=luciferase
                     /note="synthetic luc2 version of the luciferase gene"
                     /translation="MEDAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDA
                     HIEVDITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAP
                     ANDIYNERELLNSMGISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQS
                     MYTFVTSHLPPGFNEYDFVPESFDRDKTIALIMNSSGSTGLPKGVALPHRTACVRFSHA
                     RDPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYK
                     IQSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGY
                     GLTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLDTGKTLGVNQRGELCVRGPMIMS
                     GYVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESIL
                     LQHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVV
                     FVDEVPKGLTGKLDARKIREILIKAKKGGKIAV"
     polyA_signal    3680..3801
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3691..3710)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     protein_bind    complement(3916..3949)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        3959..4006
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             4038..4832
                     /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(4092..4111)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4702..4721
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     rep_origin      complement(4990..5578)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5070..5089)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    5704..5752
                     /note="synthetic polyadenylation signal"
     CDS             complement(5776..6174)
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     regulatory      6171..6180
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(6205..6562)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      6219..6354
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     enhancer        6371..6562
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     6519..6539
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
ORIGIN
        1 aagacgcgtt aattaagacg tcggcagtga aaaaaatgct ttatttgtga aatttgtgat
       61 gctattgctt tatttgtaac cattataagc tgcaataaac aagttaacaa caacaattgc
      121 attcatttta tgtttcaggt tcagggggag gtgtgggagg ttttttaaag caagtaaaac
      181 ctctacaaat gtggtatggc tgattatgat ccgcggccgc aatacgtcga cgttatcagc
      241 tgacttcgta cgagagccta ggattatggc gcgccactag ttattaatag taatcaatta
      301 cggggtcatt agttcatagc ccatatatgg agttccgcgt tacataactt acggtaaatg
      361 gcccgcctgg ctgaccgccc aacgaccccc gcccattgac gtcaataatg acgtatgttc
      421 ccatagtaac gccaataggg actttccatt gacgtcaatg ggtggagtat ttacggtaaa
      481 ctgcccactt ggcagtacat caagtgtatc atatgccaag tccgccccct attgacgtca
      541 atgacggtaa atggcccgcc tggcattatg cccagtacat gaccttacgg gactttccta
      601 cttggcagta catctacgta ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt
      661 acaccaatgg gcgtggatag cggtttgact cacggggatt tccaagtctc caccccattg
      721 acgtcaatgg gagtttgttt tggcaccaaa atcaacggga ctttccaaaa tgtcgtaata
      781 accccgcccc gttgacgcaa atgggcggta ggcgtgtacg gtgggaggtc tatataagca
      841 gagctggttt agtgaaccgt cagatcacta gaatctttat tgcggtagtt tatcacagtt
      901 aaattgctaa cgcagtcagt gcttctgaca caacagtctc gaacttaagc tgcagaagtt
      961 ggtcgtgagg cactgggcag gtaagtatca aggttacaag acaggtttaa ggagaccaat
     1021 agaaactggg cttgtcgaga cagagaagac tcttgcgttt ctgataggca cctattggtc
     1081 ttactgacat ccactttgcc tttctctcca caggtgtcca ctcccagttc aattacagct
     1141 cttaaggcta gagtattaat acgactcact ataggggcga tcgcaacaag cttcgggaag
     1201 atctggagct agcggcgggg gatccggtgg gaattctggc tcgagcggtg ggagctccgg
     1261 tgtgagcggc tggcggctgt tcaagaagat tagctgaccg accaagcgac gcccaacctg
     1321 ccatcacgga ccacaacggt ttccctctag cgggatcaat tccgcccctc tccctccctc
     1381 ccccctaacg ttactggccg aagccgcttg gaataaggcc ggtgtgcgtt tgtctatatg
     1441 ttattttcca ccatattgcc gtcttttggc aatgtgaggg cccggaaacc tggccctgtc
     1501 ttcttgacga gcattcctag gggtctttcc cctctcgcca aaggaatgca aggtctgttg
     1561 aatgtcgtga aggaagcagt tcctctggaa gcttcttgaa gacaaacaac gtctgtagcg
     1621 accctttgca ggcagcggaa ccccccacct ggcgacaggt gcctctgcgg ccaaaagcca
     1681 cgtgtataag atacacctgc aaaggcggca caaccccagt gccacgttgt gagttggata
     1741 gttgtggaaa gagtcaaatg gctctcctca agcgtattca acaaggggct gaaggatgcc
     1801 cagaaggtac cccattgtat gggatctgat ctggggcctc ggtgcacatg ctttacatgt
     1861 gtttagtcga ggttaaaaaa cgtctaggcc ccccgaacca cggggacgtg gttttccttt
     1921 gaaaaacacg atgataatat ggaagatgcc aaaaacatta agaagggccc agcgccattc
     1981 tacccactcg aagacgggac cgccggcgag cagctgcaca aagccatgaa gcgctacgcc
     2041 ctggtgcccg gcaccatcgc ctttaccgac gcacatatcg aggtggacat tacctacgcc
     2101 gagtacttcg agatgagcgt tcggctggca gaagctatga agcgctatgg gctgaataca
     2161 aaccatcgga tcgtggtgtg cagcgagaat agcttgcagt tcttcatgcc cgtgttgggt
     2221 gccctgttca tcggtgtggc tgtggcccca gctaacgaca tctacaacga gcgcgagctg
     2281 ctgaacagca tgggcatcag ccagcccacc gtcgtattcg tgagcaagaa agggctgcaa
     2341 aagatcctca acgtgcaaaa gaagctaccg atcatacaaa agatcatcat catggatagc
     2401 aagaccgact accagggctt ccaaagcatg tacaccttcg tgacttccca tttgccaccc
     2461 ggcttcaacg agtacgactt cgtgcccgag agcttcgacc gggacaaaac catcgccctg
     2521 atcatgaaca gtagtggcag taccggattg cccaagggcg tagccctacc gcaccgcacc
     2581 gcttgtgtcc gattcagtca tgcccgcgac cccatcttcg gcaaccagat catccccgac
     2641 accgctatcc tcagcgtggt gccatttcac cacggcttcg gcatgttcac cacgctgggc
     2701 tacttgatct gcggctttcg ggtcgtgctc atgtaccgct tcgaggagga gctattcttg
     2761 cgcagcttgc aagactataa gattcaatct gccctgctgg tgcccacact atttagcttc
     2821 ttcgctaaga gcactctcat cgacaagtac gacctaagca acttgcacga gatcgccagc
     2881 ggcggggcgc cgctcagcaa ggaggtaggt gaggccgtgg ccaaacgctt ccacctacca
     2941 ggcatccgcc agggctacgg cctgacagaa acaaccagcg ccattctgat cacccccgaa
     3001 ggggacgaca agcctggcgc agtaggcaag gtggtgccct tcttcgaggc taaggtggtg
     3061 gacttggaca ccggtaagac actgggtgtg aaccagcgcg gcgagctgtg cgtccgtggc
     3121 cccatgatca tgagcggcta cgttaacaac cccgaggcta caaacgctct catcgacaag
     3181 gacggctggc tgcacagcgg cgacatcgcc tactgggacg aggacgagca cttcttcatc
     3241 gtggaccggc tgaagagcct gatcaaatac aagggctacc aggtagcccc agccgaactg
     3301 gagagcatcc tgctgcaaca ccccaacatc ttcgacgccg gggtcgccgg cctgcccgac
     3361 gacgatgccg gcgagctgcc cgccgcagtc gtcgtgctgg aacacggtaa aaccatgacc
     3421 gagaaggaga tcgtggacta tgtggccagc caggttacaa ccgccaagaa gctgcgcggt
     3481 ggtgttgtgt tcgtggacga ggtgcctaaa ggactgaccg gcaagttgga cgcccgcaag
     3541 atccgcgaga ttctcattaa ggccaagaag ggcggcaaga tcgccgtgta agtttaaacg
     3601 gccgcgactc tagactgcag gcatgcaagc tgatttggct gctgccaccg ctgagcaagg
     3661 ccgcttcgag cagacatgat aagatacatt gatgagtttg gacaaaccac aactagaatg
     3721 cagtgaaaaa aatgctttat ttgtgaaatt tgtgatgcta ttgctttatt tgtaaccatt
     3781 ataagctgca ataaacaagt taacaacaac aattgcattc attttatgtt tcaggttcag
     3841 ggggagatgt gggaggtttt tttaagcaag taaaacctct acaaatgtgg taaaatctta
     3901 attaagactt ctaatataac ttcgtatagc atacattata cgaagttatg gttccactgt
     3961 tgacaattaa tcatcggcat agtatatcgg catagtataa tacgacaagg tgaggaacta
     4021 aacccaggag gcagatcatg attgaacaag atggattgca cgcaggttct ccggccgctt
     4081 gggtggagag gctattcggc tatgactggg cacaacagac aatcggctgc tctgatgccg
     4141 ccgtgttccg gctgtcagcg caggggcgcc cggttctttt tgtcaagacc gacctgtccg
     4201 gtgccctgaa tgaactgcag gacgaggcag cgcggctatc gtggctggcc acgacgggcg
     4261 ttccttgcgc agctgtgctc gacgttgtca ctgaagcggg aagggactgg ctgctattgg
     4321 gcgaagtgcc ggggcaggat ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca
     4381 tcatggctga tgcaatgcgg cggctgcata cgcttgatcc ggctacctgc ccattcgacc
     4441 accaagcgaa acatcgcatc gagcgagcac gtactcggat ggaagccggt cttgtcgatc
     4501 aggatgatct ggacgaagag catcaggggc tcgcgccagc cgaactgttc gccaggctca
     4561 aggcgcgcat gcccgacggc gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga
     4621 atatcatggt ggaaaatggc cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg
     4681 cggaccgcta tcaggacata gcgttggcta cccgtgatat tgctgaagag cttggcggcg
     4741 aatgggctga ccgcttcctc gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg
     4801 ccttctatcg ccttcttgac gagttcttct gattcgaaat gaccgaccaa gcgacgccca
     4861 accggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc
     4921 aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt
     4981 gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag
     5041 tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc
     5101 cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc
     5161 ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt
     5221 cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt
     5281 atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc
     5341 agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa
     5401 gtggtggcct aactacggct acactagaag gacagtattt ggtatctgcg ctctgctgaa
     5461 gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg
     5521 tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatttcaaga
     5581 agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg
     5641 gattttggtc atgagattat caaaaaggat cttcacctag atccttttat agtccggaat
     5701 cgtcacacaa aaaaccaaca cacagatgta atgaaaataa agatatttta ttgcggccat
     5761 cgtgatggct agagtttagc cctcccacac ataaccagag ggcagcaatt cacgaatccc
     5821 aactgccgtc ggctgtccat cactgtcctt cactatggct ttgatcccag gatgcagatc
     5881 gagaagcacc tgtcggcacc gtccgcaggg gctcaagatg cccctgttct catttccgat
     5941 cgcgacgata caagtcaggt tgccagctgc cgcagcagca gcagtgccca gcaccacgag
     6001 ttctgcacaa ggtcccccag taaaatgata tacattgaca ccagtgaaga tgcggccgtc
     6061 gctagagaga gctgcgctgg cgacgctgta gtcttcagag atggggatgc tgttgattgt
     6121 agccgttgct ctttcaatga gggtggattc ttcttgagac aaaggcttgg ccatggtggc
     6181 gctagtgtca gaagaatcga gctttttgca aaagcctagg cctccaaaaa agcctcctca
     6241 ctacttctgg aatagctcag aggccgaggc ggcctcggcc tctgcataaa taaaaaaaat
     6301 tagtcagcca tggggcggag aatgggcgga actgggcgga gttaggggcg ggatgggcgg
     6361 agttaggggc gggactatgg ttgctgacta attgagatgc atgctttgca tacttctgcc
     6421 tgctggggag cctggggact ttccacacct ggttgctgac taattgagat gcatgctttg
     6481 catacttctg cctgctgggg agcctgggga ctttccacac cctaactgac acacattcca
     6541 cagctggttc tttccgcctc agaaggtacc taaccaagtt cctctttcag aggttatttc
     6601 aggccatggt gctgcgc
//
