LOCUS       sequence-237303-        5466 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pMVP L3-L2 entry plasmid, contains polyA + CMV-eGFP-P2A-TETa-polyA 
            for 3- or 4-component MultiSite Gateway Pro assembly. Adds 
            CMV-driven GFP-P2A-TETa downstream of gene of interest..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5466)
  AUTHORS   Haldeman JM, Conway AE, Arlotto ME, Slentz DH, Muoio DM, Becker TC, 
            Newgard CB
  TITLE     Creation of versatile cloning platforms for transgene expression and
            dCas9-based epigenome editing.
  JOURNAL   Nucleic Acids Res. 2018 Dec 27. pii: 5264291. doi: 
            10.1093/nar/gky1286.
  PUBMED    30590691
REFERENCE   2  (bases 1 to 5466)
  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: "Nucleic 
            Acids Res. 2018 Dec 27. pii: 5264291. doi: 10.1093/nar/gky1286."
COMMENT     Sequence Label: IR904: pMVP (L3-L2) pA; CMV::eGFP-P2A-TETa-pA
FEATURES             Location/Qualifiers
     source          1..5466
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     92..109
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(268..295)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(387..473)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     522..544
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     536..553
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     537..553
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    570..665
                     /gene="mutant version of attL"
                     /label=attL3
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     polyA_signal    820..901
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     enhancer        965..1268
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1269..1472
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1423..1443
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             join(1538..1540,1541..1543,1544..2254)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1538..1540,1541..1543,1544..2254)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1844..1863)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2191..2212
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2270..2326
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             join(2336..2953,2954..2959,2960..2995,2996..2998,2999..3034,
                     3035..3037,3038..3073,3074..3082)
                     /codon_start=1
                     /product="improved tetracycline-controlled transactivator"
                     /label=rtTA-Advanced
                     /note="In the Tet-On(R) system, rtTA-Advanced binds to the 
                     Tet-responsive element and stimulates transcription only in
                     the presence of tetracycline or doxycycline."
                     /translation="MSRLDKSKVINGALELLNGVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALPIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEEQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDRQGAEPAFLFGLELIICGLEKQLKCESGGPADALDDFDLDMLPA
                     DALDDFDLDMLPADALDDFDLDMLPG"
     polyA_signal    3105..3226
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3116..3135)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     protein_bind    complement(3578..3677)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(3707..3726)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3708..3726)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3731..3747)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3731..3747)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3849..3868)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             3860..4669
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(3927..3946)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      4816..5404
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5305..5324
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga
       61 taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga
      121 gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca
      181 cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaata cgcgtaccgc
      241 tagccaggaa gagtttgtag aaacgcaaaa aggccatccg tcaggatggc cttctgctta
      301 gtttgatgcc tggcagttta tggcgggcgt cctgcccgcc accctccggg ccgttgcttc
      361 acaacgttca aatccgctcc cggcggattt gtcctactca ggagagcgtt caccgacaaa
      421 caacagataa aacgaaaggc ccagtcttcc gactgagcct ttcgttttat ttgatgcctg
      481 gcagttccct actctcgcgt taacgctagc atggatgttt tcccagtcac gacgttgtaa
      541 aacgacggcc agtcttaagc tcgggcccca aataatgatt ttattttgac tgatagtgac
      601 ctgttcgttg caacaaattg atgagcaatg cttttttata atgccaactt tgtataataa
      661 agttgagatc aacaaagctt cctagctcga tttaaataga taactgacta gaggatcata
      721 atcagccata ccacatttgt agaggtttta cttgctttaa aaaacctccc acacctcccc
      781 ctgaacctga aacataaaat gaatgcaatt gttgttgtta acttgtttat tgcagcttat
      841 aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg
      901 cctcgagtct agaatagtaa tcaattacgg ggtcattagt tcatagccca tatatggagt
      961 tccgcgttac ataacttacg gtaaatggcc cgcctggctg accgcccaac gacccccgcc
     1021 cattgacgtc aataatgacg tatgttccca tagtaacgcc aatagggact ttccattgac
     1081 gtcaatgggt ggagtattta cggtaaactg cccacttggc agtacatcaa gtgtatcata
     1141 tgccaagtac gccccctatt gacgtcaatg acggtaaatg gcccgcctgg cattatgccc
     1201 agtacatgac cttatgggac tttcctactt ggcagtacat ctacgtatta gtcatcgcta
     1261 ttaccatggt gatgcggttt tggcagtaca tcaatgggcg tggatagcgg tttgactcac
     1321 ggggatttcc aagtctccac cccattgacg tcaatgggag tttgttttgg caccaaaatc
     1381 aacgggactt tccaaaatgt cgtaacaact ccgccccatt gacgcaaatg ggcggtaggc
     1441 gtgtacggtg ggaggtctat ataagcagag ctggtttagt gaaccgtcag atccgctagc
     1501 gctaccggac tcagatctcg aattcgactg gcgcgccatg gtgagcaagg gcgaggagct
     1561 gttcaccggg gtggtgccca tcctggtcga gctggacggc gacgtaaacg gccacaagtt
     1621 cagcgtgtcc ggcgagggcg agggcgatgc cacctacggc aagctgaccc tgaagttcat
     1681 ctgcaccacc ggcaagctgc ccgtgccctg gcccaccctc gtgaccaccc tgacctacgg
     1741 cgtgcagtgc ttcagccgct accccgacca catgaagcag cacgacttct tcaagtccgc
     1801 catgcccgaa ggctacgtcc aggagcgcac catcttcttc aaggacgacg gcaactacaa
     1861 gacccgcgcc gaggtgaagt tcgagggcga caccctggtg aaccgcatcg agctgaaggg
     1921 catcgacttc aaggaggacg gcaacatcct ggggcacaag ctggagtaca actacaacag
     1981 ccacaacgtc tatatcatgg ccgacaagca gaagaacggc atcaaggtga acttcaagat
     2041 ccgccacaac atcgaggacg gcagcgtgca gctcgccgac cactaccagc agaacacccc
     2101 catcggcgac ggccccgtgc tgctgcccga caaccactac ctgagcaccc agtccgccct
     2161 gagcaaagac cccaacgaga agcgcgatca catggtcctg ctggagttcg tgaccgccgc
     2221 cgggatcact ctcggcatgg acgagctgta caagcgcgcc ggatcaggag ctactaattt
     2281 ctctctgtta aagcaggcag gagatgtgga ggagaatcca ggtcctcaat tcaccatgtc
     2341 cagactggac aagagcaaag tcataaacgg cgctctggaa ttactcaatg gagtcggtat
     2401 cgaaggcctg acgacaagga aactcgctca aaagctggga gttgagcagc ctaccctgta
     2461 ctggcacgtg aagaacaagc gggccctgct cgatgccctg ccaatcgaga tgctggacag
     2521 gcatcatacc cacttctgcc ccctggaagg cgagtcatgg caagactttc tgcggaacaa
     2581 cgccaagtca ttccgctgtg ctctcctctc acatcgcgac ggggctaaag tgcatctcgg
     2641 cacccgccca acagagaaac agtacgaaac cctggaaaat cagctcgcgt tcctgtgtca
     2701 gcaaggcttc tccctggaga acgcactgta cgctctgtcc gccgtgggcc actttacact
     2761 gggctgcgta ttggaggaac aggagcatca agtagcaaaa gaggaaagag agacacctac
     2821 caccgattct atgcccccac ttctgagaca agcaattgag ctgttcgacc ggcagggagc
     2881 cgaacctgcc ttccttttcg gcctggaact aatcatatgt ggcctggaga aacagctaaa
     2941 gtgcgaaagc ggcgggccgg ccgacgccct tgacgatttt gacttagaca tgctcccagc
     3001 cgatgccctt gacgactttg accttgatat gctgcctgct gacgctcttg acgattttga
     3061 ccttgacatg ctccccgggt aactaagtaa gaatccagac atgataagat acattgatga
     3121 gtttggacaa accacaacta gaatgcagtg aaaaaaatgc tttatttgtg aaatttgtga
     3181 tgctattgct ttatttgtaa ccattataag ctgcaataaa caagttaaca acaacaattg
     3241 cattcatttt atgtttcagg ttcaggggga ggtgtgggag gttttttaaa gcaagtaaaa
     3301 cctctacaaa tgtggtatgg ctgattatga tcctgcaagc ctcgtcgtcc tggccggacc
     3361 acgctatctg tgcaaggtcc ccggccccgg acgcgcgctc catgagcaga gcgcccgccg
     3421 ccgaggcgaa gactcgggcg gcgccctgcc cgtcccacca ggtcaacagg cggtaaccgg
     3481 cctcttcatc gggaatgcgc gcgaccttca gcatcgccgg catgtccccc tggcggacgg
     3541 gaagtatcca gctcgaccag gcttgtcgac tcgaagaacc cagctttctt gtacaaagtt
     3601 ggcattataa gaaagcattg cttatcaatt tgttgcaacg aacaggtcac tatcagtcaa
     3661 aataaaatca ttatttgcca tccagctgca gggcggccgc gatatcccct atagtgagtc
     3721 gtattacatg gtcatagctg tttcctggca gctctggccc gtgtctcaaa atctctgatg
     3781 ttacattgca caagataaaa atatatcatc atgaacaata aaactgtctg cttacataaa
     3841 cagtaataca aggggtgtta tgagccatat tcaacgggaa acgtcgaggc cgcgattaaa
     3901 ttccaacatg gatgctgatt tatatgggta taaatgggct cgcgataatg tcgggcaatc
     3961 aggtgcgaca atctatcgct tgtatgggaa gcccgatgcg ccagagttgt ttctgaaaca
     4021 tggcaaaggt agcgttgcca atgatgttac agatgagatg gtcagactaa actggctgac
     4081 ggaatttatg cctcttccga ccatcaagca ttttatccgt actcctgatg atgcatggtt
     4141 actcaccact gcgatccccg gaaaaacagc attccaggta ttagaagaat atcctgattc
     4201 aggtgaaaat attgttgatg cgctggcagt gttcctgcgc cggttgcatt cgattcctgt
     4261 ttgtaattgt ccttttaaca gcgatcgcgt atttcgtctc gctcaggcgc aatcacgaat
     4321 gaataacggt ttggttgatg cgagtgattt tgatgacgag cgtaatggct ggcctgttga
     4381 acaagtctgg aaagaaatgc ataaactttt gccattctca ccggattcag tcgtcactca
     4441 tggtgatttc tcacttgata accttatttt tgacgagggg aaattaatag gttgtattga
     4501 tgttggacga gtcggaatcg cagaccgata ccaggatctt gccatcctat ggaactgcct
     4561 cggtgagttt tctccttcat tacagaaacg gctttttcaa aaatatggta ttgataatcc
     4621 tgatatgaat aaattgcagt ttcatttgat gctcgatgag tttttctaat cagaattggt
     4681 taattggttg taacactggc agagcattac gctgacttga cgggacggcg caagctcatg
     4741 accaaaatcc cttaacgtga gttacgcgtc gttccactga gcgtcagacc ccgtagaaaa
     4801 gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa
     4861 aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc
     4921 gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag tgtagccgta
     4981 gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct
     5041 gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg
     5101 atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag
     5161 cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc
     5221 cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg
     5281 agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt
     5341 tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg
     5401 gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca
     5461 catgtt
//
