LOCUS       Exported                3415 bp ds-DNA     circular SYN 13-MAY-2021
DEFINITION  pMVP L5-L4 entry plasmid, contains eCFP-P2A for 4-component 
            MultiSite Gateway Pro assembly. Allows expression of N-term eCFP 
            linked by P2A to gene of interest..
ACCESSION   .
VERSION     .
KEYWORDS    KP401: pMVP (L5-L4) eCFP-P2A
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3415)
  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 3415)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 13, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..3415
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     92..109
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      268..295
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      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=attL5
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     5'UTR           680..722
                     /label=Xenopus globin 5'-UTR
                     /note="translational enhancer from the 5'-UTR of the major 
                     beta-globin gene of Xenopus laevis"
     CDS             734..1450
                     /codon_start=1
                     /product="enhanced CFP"
                     /label=ECFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTWGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYISHNVYITADKQKNGIK
                     ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(779..800)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1040..1059)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1387..1408
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1463..1519
                     /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"
     protein_bind    complement(1530..1625)
                     /gene="mutant version of attL"
                     /label=attL4
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(1656..1675)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1657..1675)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1680..1696)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1680..1696)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1798..1817)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             1809..2618
                     /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(1876..1895)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      2765..3353
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3254..3273
                     /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 tgtatacaaa
      661 agttgtagaa tacaagcttc ttgttctttt tgcagaagct cagaataaac gctcaacttt
      721 ggcagatctg aacatggtga gcaagggcga ggagctgttc accggggtgg tgcccatcct
      781 ggtcgagctg gacggcgacg taaacggcca caagttcagc gtgtccggcg agggcgaggg
      841 cgatgccacc tacggcaagc tgaccctgaa gttcatctgc accaccggca agctgcccgt
      901 gccctggccc accctcgtga ccaccctgac ctggggcgtg cagtgcttca gccgctaccc
      961 cgaccacatg aagcagcacg acttcttcaa gtccgccatg cccgaaggct acgtccagga
     1021 gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc cgcgccgagg tgaagttcga
     1081 gggcgacacc ctggtgaacc gcatcgagct gaagggcatc gacttcaagg aggacggcaa
     1141 catcctgggg cacaagctgg agtacaacta catcagccac aacgtctata tcaccgccga
     1201 caagcagaag aacggcatca aggccaactt caagatccgc cacaacatcg aggacggcag
     1261 cgtgcagctc gccgaccact accagcagaa cacccccatc ggcgacggcc ccgtgctgct
     1321 gcccgacaac cactacctga gcacccagtc cgccctgagc aaagacccca acgagaagcg
     1381 cgatcacatg gtcctgctgg agttcgtgac cgccgccggg atcactctcg gcatggacga
     1441 gctgtacaag gccggatccg gagctactaa tttctctctg ttaaagcagg caggagatgt
     1501 ggaggagaat ccaggtccta aggcccaccc aacttttcta tacaaagttg gcattataag
     1561 aaagcattgc ttatcaattt gttgcaacga acaggtcact atcagtcaaa ataaaatcat
     1621 tatttgccat ccagctgcag ggcggccgcg atatccccta tagtgagtcg tattacatgg
     1681 tcatagctgt ttcctggcag ctctggcccg tgtctcaaaa tctctgatgt tacattgcac
     1741 aagataaaaa tatatcatca tgaacaataa aactgtctgc ttacataaac agtaatacaa
     1801 ggggtgttat gagccatatt caacgggaaa cgtcgaggcc gcgattaaat tccaacatgg
     1861 atgctgattt atatgggtat aaatgggctc gcgataatgt cgggcaatca ggtgcgacaa
     1921 tctatcgctt gtatgggaag cccgatgcgc cagagttgtt tctgaaacat ggcaaaggta
     1981 gcgttgccaa tgatgttaca gatgagatgg tcagactaaa ctggctgacg gaatttatgc
     2041 ctcttccgac catcaagcat tttatccgta ctcctgatga tgcatggtta ctcaccactg
     2101 cgatccccgg aaaaacagca ttccaggtat tagaagaata tcctgattca ggtgaaaata
     2161 ttgttgatgc gctggcagtg ttcctgcgcc ggttgcattc gattcctgtt tgtaattgtc
     2221 cttttaacag cgatcgcgta tttcgtctcg ctcaggcgca atcacgaatg aataacggtt
     2281 tggttgatgc gagtgatttt gatgacgagc gtaatggctg gcctgttgaa caagtctgga
     2341 aagaaatgca taaacttttg ccattctcac cggattcagt cgtcactcat ggtgatttct
     2401 cacttgataa ccttattttt gacgagggga aattaatagg ttgtattgat gttggacgag
     2461 tcggaatcgc agaccgatac caggatcttg ccatcctatg gaactgcctc ggtgagtttt
     2521 ctccttcatt acagaaacgg ctttttcaaa aatatggtat tgataatcct gatatgaata
     2581 aattgcagtt tcatttgatg ctcgatgagt ttttctaatc agaattggtt aattggttgt
     2641 aacactggca gagcattacg ctgacttgac gggacggcgc aagctcatga ccaaaatccc
     2701 ttaacgtgag ttacgcgtcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat
     2761 cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc
     2821 taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg
     2881 gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc
     2941 acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg
     3001 ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg
     3061 ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa
     3121 cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg
     3181 aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga
     3241 gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct
     3301 gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca
     3361 gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgtt
//
