LOCUS       sequence-471889-        4613 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express SmBiT-CRAF RBD 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 4613)
  AUTHORS   Promega
  TITLE     Promega Plasmid Collection
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 4613)
  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: 
            "Unpublished"
COMMENT     Sequence Label: SmBiT-CRAF RBD BiBiT-Ready Vector
FEATURES             Location/Qualifiers
     source          1..4613
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(747..1541)
                     /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(858..877)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     1468..1487
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(1573..1620)
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     protein_bind    1630..1663
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     polyA_signal    1778..1899
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     1869..1888
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(2399..2418)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2400..2418)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2460..2479)
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     intron          2462..2594
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     promoter        complement(2730..2933)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(2759..2779)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        2934..3237
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     polyA_signal    3470..3551
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     promoter        3630..3987
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3630..3821
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3653..3673)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3838..3973
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     regulatory      4012..4021
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             4018..4416
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     polyA_signal    4440..4488
                     /note="synthetic polyadenylation signal"
ORIGIN
        1 ttgaaatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgtcct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
      661 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccggttg
      721 ggcgtcgctt ggtcggtcat ttcgaatcag aagaactcgt caagaaggcg atagaaggcg
      781 atgcgctgcg aatcgggagc ggcgataccg taaagcacga ggaagcggtc agcccattcg
      841 ccgccaagct cttcagcaat atcacgggta gccaacgcta tgtcctgata gcggtccgcc
      901 acacccagcc ggccacagtc gatgaatcca gaaaagcggc cattttccac catgatattc
      961 ggcaagcagg catcgccatg ggtcacgacg agatcctcgc cgtcgggcat gcgcgccttg
     1021 agcctggcga acagttcggc tggcgcgagc ccctgatgct cttcgtccag atcatcctga
     1081 tcgacaagac cggcttccat ccgagtacgt gctcgctcga tgcgatgttt cgcttggtgg
     1141 tcgaatgggc aggtagccgg atcaagcgta tgcagccgcc gcattgcatc agccatgatg
     1201 gatactttct cggcaggagc aaggtgagat gacaggagat cctgccccgg cacttcgccc
     1261 aatagcagcc agtcccttcc cgcttcagtg acaacgtcga gcacagctgc gcaaggaacg
     1321 cccgtcgtgg ccagccacga tagccgcgct gcctcgtcct gcagttcatt cagggcaccg
     1381 gacaggtcgg tcttgacaaa aagaaccggg cgcccctgcg ctgacagccg gaacacggcg
     1441 gcatcagagc agccgattgt ctgttgtgcc cagtcatagc cgaatagcct ctccacccaa
     1501 gcggccggag aacctgcgtg caatccatct tgttcaatca tgatctgcct cctgggttta
     1561 gttcctcacc ttgtcgtatt atactatgcc gatatactat gccgatgatt aattgtcaac
     1621 agtggatcca taacttcgta taatgtatgc tatacgaagt tatattagaa ttcttaatta
     1681 agattttacc acatttgtag aggttttact tgcttaaaaa aacctcccac atctccccct
     1741 gaacctgaaa cataaaatga atgcaattgt tgttgttaac ttgtttattg cagcttataa
     1801 tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca
     1861 ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgct cgaagcggcc
     1921 ttgctcagcg gtggcagcag ccaactcagc ttcctttcgg gctttgttag cagccggatc
     1981 agcttgcatg cctgcaggtc gactctagag gatccccggg taccgagccc gaattcgttt
     2041 aaacatgatc caggaaatct acttgaagtt cttctccaat caaagacgca gcatcagtat
     2101 tccaatctaa gcgtgctttt ttacctttgt gttcgtggag aagtctgaac actgcacagc
     2161 actctggttg caggcccctc accttgagtg ctttcataag gcagtcatgc aagctcattc
     2221 catttcgcac attgaccact gttctttgct tgttcggcaa gaaaacacgg atagtgttgc
     2281 ttgtcttaga aggcatggcg atcgcaccgc tcgagcctcc acctccgctc ccgccaccac
     2341 cggagctccc gagaatctcc tcgaacagcc ggtagccggt caccatggtg agcgctagcc
     2401 ctatagtgag tcgtattaat actctagcct taagagctgt aattgaactg ggagtggaca
     2461 cctgtggaga gaaaggcaaa gtggatgtca gtaagaccaa taggtgccta tcagaaacgc
     2521 aagagtcttc tctgtctcga caagcccagt ttctattggt ctccttaaac ctgtcttgta
     2581 accttgatac ttacctgccc agtgcctcac gaccaacttc tgcagcttaa gttcgagact
     2641 gttgtgtcag aagcactgac tgcgttagca atttaactgt gataaactac cgcaataaag
     2701 cttctagtga tctgacggtt cactaaacca gctctgctta tatagacctc ccaccgtaca
     2761 cgcctaccgc ccatttgcgt caacggggcg gggttattac gacattttgg aaagtcccgt
     2821 tgattttggt gccaaaacaa actcccattg acgtcaatgg ggtggagact tggaaatccc
     2881 cgtgagtcaa accgctatcc acgcccattg gtgtactgcc aaaaccgcat caccatggta
     2941 atagcgatga ctaatacgta gatgtactgc caagtaggaa agtcccgtaa ggtcatgtac
     3001 tgggcataat gccaggcggg ccatttaccg tcattgacgt caataggggg cggacttggc
     3061 atatgataca cttgatgtac tgccaagtgg gcagtttacc gtaaatactc cacccattga
     3121 cgtcaatgga aagtccctat tggcgttact atgggaacat acgtcattat tgacgtcaat
     3181 gggcgggggt cgttgggcgg tcagccaggc gggccattta ccgtaagtta tgtaacgcgg
     3241 aactccatat atgggctatg aactaatgac cccgtaattg attactatta ataactagtg
     3301 gcgcgccata atcctaggct ctcgtacgaa gtcagctgat aacgtcgacg tattgcggcc
     3361 gcggatcata atcagccata ccacatttgt agaggtttta cttgctttaa aaaacctccc
     3421 acacctcccc ctgaacctga aacataaaat gaatgcaatt gttgttgtta acttgtttat
     3481 tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt
     3541 tttttcactg ccgacgtctt aattaacgcg tcttgcgcag caccatggcc tgaaataacc
     3601 tctgaaagag gaacttggtt aggtaccttc tgaggcggaa agaaccagct gtggaatgtg
     3661 tgtcagttag ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg
     3721 catctcaatt agtcagcaac caggtgtgga aagtccccag gctccccagc aggcagaagt
     3781 atgcaaagca tgcatctcaa ttagtcagca accatagtcc cgcccctaac tccgcccatc
     3841 ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt
     3901 atttatgcag aggccgaggc cgcctcggcc tctgagctat tccagaagta gtgaggaggc
     3961 ttttttggag gcctaggctt ttgcaaaaag ctcgattctt ctgacactag cgccaccatg
     4021 gccaagcctt tgtctcaaga agaatccacc ctcattgaaa gagcaacggc tacaatcaac
     4081 agcatcccca tctctgaaga ctacagcgtc gccagcgcag ctctctctag cgacggccgc
     4141 atcttcactg gtgtcaatgt atatcatttt actgggggac cttgtgcaga actcgtggtg
     4201 ctgggcactg ctgctgctgc ggcagctggc aacctgactt gtatcgtcgc gatcggaaat
     4261 gagaacaggg gcatcttgag cccctgcgga cggtgccgac aggtgcttct cgatctgcat
     4321 cctgggatca aagccatagt gaaggacagt gatggacagc cgacggcagt tgggattcgt
     4381 gaattgctgc cctctggtta tgtgtgggag ggctaaactc tagccatcac gatggccgca
     4441 ataaaatatc tttattttca ttacatctgt gtgttggttt tttgtgtgac gattccggac
     4501 tataaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt
     4561 gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttc
//
