LOCUS       sequence-237278-        3926 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pMVP L3-L2 entry plasmid, contains Hygro-P2A + polyA for 3- or 
            4-component MultiSite Gateway Pro assembly. Allows expression of 
            C-term Hygro selection marker linked by P2A to gene of interest..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3926)
  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 3926)
  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: KZ201: pMVP (L3-L2) P2A-Hygro + polyA
FEATURES             Location/Qualifiers
     source          1..3926
                     /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"
     CDS             680..736
                     /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             743..1780
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     EVGRV"
     RBS             1759..1767
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     primer_bind     complement(1816..1833)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1822..2029
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     protein_bind    complement(2038..2137)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(2167..2186)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2168..2186)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2191..2207)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2191..2207)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2309..2328)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             2320..3129
                     /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(2387..2406)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      3276..3864
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3765..3784
                     /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 agttggtggc ggatccggag ctactaattt ctctctgtta aagcaggcag gagatgtgga
      721 ggagaatcca ggtcctaagg gaatgaagaa gcccgaactc accgctacca gcgttgaaaa
      781 atttctcatc gagaagttcg acagtgtgag cgacctgatg cagttgtcgg agggcgaaga
      841 gagccgagcc ttcagcttcg atgtcggcgg acgcggctat gtactgcggg tgaatagctg
      901 cgctgatggc ttctacaaag accgctacgt gtaccgccac ttcgccagcg ctgcactacc
      961 catccccgaa gtgttggaca tcggcgagtt cagcgagagc ctgacatact gcatcagtag
     1021 acgcgcccaa ggcgttactc tccaagacct ccccgaaaca gagctgcctg ctgtgttaca
     1081 gcctgtcgcc gaagctatgg atgctattgc cgccgccgac ctcagtcaaa ccagcggctt
     1141 cggcccattc gggccccaag gcatcggcca gtacacaacc tggcgggatt tcatttgcgc
     1201 cattgctgat ccccatgtct accactggca gaccgtgatg gacgacaccg tgtccgccag
     1261 cgtagctcaa gccctggacg aactgatgct gtgggccgaa gactgtcccg aggtgcgcca
     1321 cctcgtccat gccgacttcg gcagcaacaa cgtcctgacc gacaacggcc gcatcaccgc
     1381 cgtaatcgac tggtccgaag ctatgttcgg ggacagtcag tacgaggtgg ccaacatctt
     1441 cttctggcgg ccctggctgg cttgcatgga gcagcagact cgctacttcg agcgccggca
     1501 tcccgagctg gccggcagcc ctcgtctgcg agcctacatg ctgcgcatcg gcctggatca
     1561 gctctaccag agcctcgtgg acggcaactt cgacgatgct gcctgggctc aaggccgctg
     1621 cgatgccatc gtccgcagcg gggccggcac cgtcggtcgc acacaaatcg ctcgccggag
     1681 cgcagccgta tggaccgacg gctgcgtcga ggtgctggcc gacagcggca accgccggcc
     1741 cagtacacga ccgcgcgcta aggaggtagg tcgagtttaa gaattctgaa gcttagccta
     1801 gagctcgctg atcagcctcg actgtgcctt ctagttgcca gccatctgtt gtttgcccct
     1861 cccccgtgcc ttccttgacc ctggaaggtg ccactcccac tgtcctttcc taataaaatg
     1921 aggaaattgc atcgcattgt ctgagtaggt gtcattctat tctggggggt ggggtggggc
     1981 aggacagcaa gggggaggat tgggaagaga atagcaggca tgctggggag gtacctgacc
     2041 cagctttctt gtacaaagtt ggcattataa gaaagcattg cttatcaatt tgttgcaacg
     2101 aacaggtcac tatcagtcaa aataaaatca ttatttgcca tccagctgca gggcggccgc
     2161 gatatcccct atagtgagtc gtattacatg gtcatagctg tttcctggca gctctggccc
     2221 gtgtctcaaa atctctgatg ttacattgca caagataaaa atatatcatc atgaacaata
     2281 aaactgtctg cttacataaa cagtaataca aggggtgtta tgagccatat tcaacgggaa
     2341 acgtcgaggc cgcgattaaa ttccaacatg gatgctgatt tatatgggta taaatgggct
     2401 cgcgataatg tcgggcaatc aggtgcgaca atctatcgct tgtatgggaa gcccgatgcg
     2461 ccagagttgt ttctgaaaca tggcaaaggt agcgttgcca atgatgttac agatgagatg
     2521 gtcagactaa actggctgac ggaatttatg cctcttccga ccatcaagca ttttatccgt
     2581 actcctgatg atgcatggtt actcaccact gcgatccccg gaaaaacagc attccaggta
     2641 ttagaagaat atcctgattc aggtgaaaat attgttgatg cgctggcagt gttcctgcgc
     2701 cggttgcatt cgattcctgt ttgtaattgt ccttttaaca gcgatcgcgt atttcgtctc
     2761 gctcaggcgc aatcacgaat gaataacggt ttggttgatg cgagtgattt tgatgacgag
     2821 cgtaatggct ggcctgttga acaagtctgg aaagaaatgc ataaactttt gccattctca
     2881 ccggattcag tcgtcactca tggtgatttc tcacttgata accttatttt tgacgagggg
     2941 aaattaatag gttgtattga tgttggacga gtcggaatcg cagaccgata ccaggatctt
     3001 gccatcctat ggaactgcct cggtgagttt tctccttcat tacagaaacg gctttttcaa
     3061 aaatatggta ttgataatcc tgatatgaat aaattgcagt ttcatttgat gctcgatgag
     3121 tttttctaat cagaattggt taattggttg taacactggc agagcattac gctgacttga
     3181 cgggacggcg caagctcatg accaaaatcc cttaacgtga gttacgcgtc gttccactga
     3241 gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta
     3301 atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa
     3361 gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact
     3421 gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca
     3481 tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt
     3541 accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg
     3601 ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag
     3661 cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta
     3721 agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat
     3781 ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg
     3841 tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc
     3901 ttttgctggc cttttgctca catgtt
//
