LOCUS       sequence-237316-        5290 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pMVP L3-L2 entry plasmid, contains polyA + RIP-eGFP-P2A-TETa-polyA 
            for 3- or 4-component MultiSite Gateway Pro assembly. Adds 
            RIP-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 5290)
  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 5290)
  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: IG806: pMVP (L3-L2) pA; RIP::eGFP-P2A-TETa-pA
FEATURES             Location/Qualifiers
     source          1..5290
                     /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"
     CDS             join(1362..1364,1365..1367,1368..2078)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1362..1364,1365..1367,1368..2078)
                     /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(1668..1687)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2015..2036
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2094..2150
                     /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(2160..2777,2778..2783,2784..2819,2820..2822,2823..2858,
                     2859..2861,2862..2897,2898..2906)
                     /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    2929..3050
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2940..2959)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     protein_bind    complement(3402..3501)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(3531..3550)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3532..3550)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3555..3571)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3555..3571)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3673..3692)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             3684..4493
                     /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(3751..3770)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      4640..5228
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5129..5148
                     /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 agactgagca agaatccagc tatcaataga aactatgaaa cagttccagg
      961 gacaaagata ccaggtcccc aacaactgca actttctggg aaatgaggtg gaaaatgctc
     1021 agccaaggaa aaagagggcc ttaccctctc tgggacaatg attgtgctgt gaactgcttc
     1081 atcaggccat ctggcccctt gttaataatc taattaccct aggtctaagt agagttgttg
     1141 acgtccaatg agcgctttct gcagacttag cactaggcaa gtgtttggaa attacagctt
     1201 cagcccctct cgccatctgc ctacctaccc ctcctagagc ccttaatggg ccaaacggca
     1261 aagtccaggg ggcagagagg aggtgctttg gactataaag ctagtggaga cccagtaact
     1321 cccaacccta agtgaccagc tacgaattcg actggcgcgc catggtgagc aagggcgagg
     1381 agctgttcac cggggtggtg cccatcctgg tcgagctgga cggcgacgta aacggccaca
     1441 agttcagcgt gtccggcgag ggcgagggcg atgccaccta cggcaagctg accctgaagt
     1501 tcatctgcac caccggcaag ctgcccgtgc cctggcccac cctcgtgacc accctgacct
     1561 acggcgtgca gtgcttcagc cgctaccccg accacatgaa gcagcacgac ttcttcaagt
     1621 ccgccatgcc cgaaggctac gtccaggagc gcaccatctt cttcaaggac gacggcaact
     1681 acaagacccg cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc atcgagctga
     1741 agggcatcga cttcaaggag gacggcaaca tcctggggca caagctggag tacaactaca
     1801 acagccacaa cgtctatatc atggccgaca agcagaagaa cggcatcaag gtgaacttca
     1861 agatccgcca caacatcgag gacggcagcg tgcagctcgc cgaccactac cagcagaaca
     1921 cccccatcgg cgacggcccc gtgctgctgc ccgacaacca ctacctgagc acccagtccg
     1981 ccctgagcaa agaccccaac gagaagcgcg atcacatggt cctgctggag ttcgtgaccg
     2041 ccgccgggat cactctcggc atggacgagc tgtacaagcg cgccggatca ggagctacta
     2101 atttctctct gttaaagcag gcaggagatg tggaggagaa tccaggtcct caattcacca
     2161 tgtccagact ggacaagagc aaagtcataa acggcgctct ggaattactc aatggagtcg
     2221 gtatcgaagg cctgacgaca aggaaactcg ctcaaaagct gggagttgag cagcctaccc
     2281 tgtactggca cgtgaagaac aagcgggccc tgctcgatgc cctgccaatc gagatgctgg
     2341 acaggcatca tacccacttc tgccccctgg aaggcgagtc atggcaagac tttctgcgga
     2401 acaacgccaa gtcattccgc tgtgctctcc tctcacatcg cgacggggct aaagtgcatc
     2461 tcggcacccg cccaacagag aaacagtacg aaaccctgga aaatcagctc gcgttcctgt
     2521 gtcagcaagg cttctccctg gagaacgcac tgtacgctct gtccgccgtg ggccacttta
     2581 cactgggctg cgtattggag gaacaggagc atcaagtagc aaaagaggaa agagagacac
     2641 ctaccaccga ttctatgccc ccacttctga gacaagcaat tgagctgttc gaccggcagg
     2701 gagccgaacc tgccttcctt ttcggcctgg aactaatcat atgtggcctg gagaaacagc
     2761 taaagtgcga aagcggcggg ccggccgacg cccttgacga ttttgactta gacatgctcc
     2821 cagccgatgc ccttgacgac tttgaccttg atatgctgcc tgctgacgct cttgacgatt
     2881 ttgaccttga catgctcccc gggtaactaa gtaagaatcc agacatgata agatacattg
     2941 atgagtttgg acaaaccaca actagaatgc agtgaaaaaa atgctttatt tgtgaaattt
     3001 gtgatgctat tgctttattt gtaaccatta taagctgcaa taaacaagtt aacaacaaca
     3061 attgcattca ttttatgttt caggttcagg gggaggtgtg ggaggttttt taaagcaagt
     3121 aaaacctcta caaatgtggt atggctgatt atgatcctgc aagcctcgtc gtcctggccg
     3181 gaccacgcta tctgtgcaag gtccccggcc ccggacgcgc gctccatgag cagagcgccc
     3241 gccgccgagg cgaagactcg ggcggcgccc tgcccgtccc accaggtcaa caggcggtaa
     3301 ccggcctctt catcgggaat gcgcgcgacc ttcagcatcg ccggcatgtc cccctggcgg
     3361 acgggaagta tccagctcga ccaggcttgt cgactcgaag aacccagctt tcttgtacaa
     3421 agttggcatt ataagaaagc attgcttatc aatttgttgc aacgaacagg tcactatcag
     3481 tcaaaataaa atcattattt gccatccagc tgcagggcgg ccgcgatatc ccctatagtg
     3541 agtcgtatta catggtcata gctgtttcct ggcagctctg gcccgtgtct caaaatctct
     3601 gatgttacat tgcacaagat aaaaatatat catcatgaac aataaaactg tctgcttaca
     3661 taaacagtaa tacaaggggt gttatgagcc atattcaacg ggaaacgtcg aggccgcgat
     3721 taaattccaa catggatgct gatttatatg ggtataaatg ggctcgcgat aatgtcgggc
     3781 aatcaggtgc gacaatctat cgcttgtatg ggaagcccga tgcgccagag ttgtttctga
     3841 aacatggcaa aggtagcgtt gccaatgatg ttacagatga gatggtcaga ctaaactggc
     3901 tgacggaatt tatgcctctt ccgaccatca agcattttat ccgtactcct gatgatgcat
     3961 ggttactcac cactgcgatc cccggaaaaa cagcattcca ggtattagaa gaatatcctg
     4021 attcaggtga aaatattgtt gatgcgctgg cagtgttcct gcgccggttg cattcgattc
     4081 ctgtttgtaa ttgtcctttt aacagcgatc gcgtatttcg tctcgctcag gcgcaatcac
     4141 gaatgaataa cggtttggtt gatgcgagtg attttgatga cgagcgtaat ggctggcctg
     4201 ttgaacaagt ctggaaagaa atgcataaac ttttgccatt ctcaccggat tcagtcgtca
     4261 ctcatggtga tttctcactt gataacctta tttttgacga ggggaaatta ataggttgta
     4321 ttgatgttgg acgagtcgga atcgcagacc gataccagga tcttgccatc ctatggaact
     4381 gcctcggtga gttttctcct tcattacaga aacggctttt tcaaaaatat ggtattgata
     4441 atcctgatat gaataaattg cagtttcatt tgatgctcga tgagtttttc taatcagaat
     4501 tggttaattg gttgtaacac tggcagagca ttacgctgac ttgacgggac ggcgcaagct
     4561 catgaccaaa atcccttaac gtgagttacg cgtcgttcca ctgagcgtca gaccccgtag
     4621 aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa
     4681 caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt
     4741 ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgttctt ctagtgtagc
     4801 cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa
     4861 tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa
     4921 gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc
     4981 ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa
     5041 gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa
     5101 caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg
     5161 ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc
     5221 tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg
     5281 ctcacatgtt
//
