LOCUS       Exported                4696 bp ds-DNA     circular SYN 13-MAY-2021
DEFINITION  pMVP L3-L2 entry plasmid, contains HA tag-polyA + CMV-TETa-polyA for
            3- or 4-component MultiSite Gateway Pro assembly. Adds C-term HA tag
            to gene plus downstream CMV-driven TETa.
ACCESSION   .
VERSION     .
KEYWORDS    KO602: pMVP (L3-L2) HA tag + pA; CMV::TETa-pA
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4696)
  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 4696)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 13, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..4696
                     /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=attL3
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             686..712
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     polyA_signal    845..926
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     enhancer        990..1293
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1294..1497
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1448..1468
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             1566..2312
                     /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    2335..2456
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2346..2365)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     protein_bind    complement(2808..2907)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(2937..2956)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2938..2956)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2961..2977)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2961..2977)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3079..3098)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             3090..3899
                     /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(3157..3176)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      4046..4634
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4535..4554
                     /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 aacaaagctg gatcctaccc atacgatgtg ccagattacg cttgatttaa
      721 atagataact gactagagga tcataatcag ccataccaca tttgtagagg ttttacttgc
      781 tttaaaaaac ctcccacacc tccccctgaa cctgaaacat aaaatgaatg caattgttgt
      841 tgttaacttg tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt
      901 cacaaataaa gcattttttt cactgcctcg agtctagaat agtaatcaat tacggggtca
      961 ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct
     1021 ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta
     1081 acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac
     1141 ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt
     1201 aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag
     1261 tacatctacg tattagtcat cgctattacc atggtgatgc ggttttggca gtacatcaat
     1321 gggcgtggat agcggtttga ctcacgggga tttccaagtc tccaccccat tgacgtcaat
     1381 gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa caactccgcc
     1441 ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag cagagctggt
     1501 ttagtgaacc gtcagatccg ctagcgctac cggactcaga tctcgaattc gactggcgcg
     1561 ccaccatgtc cagactggac aagagcaaag tcataaacgg cgctctggaa ttactcaatg
     1621 gagtcggtat cgaaggcctg acgacaagga aactcgctca aaagctggga gttgagcagc
     1681 ctaccctgta ctggcacgtg aagaacaagc gggccctgct cgatgccctg ccaatcgaga
     1741 tgctggacag gcatcatacc cacttctgcc ccctggaagg cgagtcatgg caagactttc
     1801 tgcggaacaa cgccaagtca ttccgctgtg ctctcctctc acatcgcgac ggggctaaag
     1861 tgcatctcgg cacccgccca acagagaaac agtacgaaac cctggaaaat cagctcgcgt
     1921 tcctgtgtca gcaaggcttc tccctggaga acgcactgta cgctctgtcc gccgtgggcc
     1981 actttacact gggctgcgta ttggaggaac aggagcatca agtagcaaaa gaggaaagag
     2041 agacacctac caccgattct atgcccccac ttctgagaca agcaattgag ctgttcgacc
     2101 ggcagggagc cgaacctgcc ttccttttcg gcctggaact aatcatatgt ggcctggaga
     2161 aacagctaaa gtgcgaaagc ggcgggccgg ccgacgccct tgacgatttt gacttagaca
     2221 tgctcccagc cgatgccctt gacgactttg accttgatat gctgcctgct gacgctcttg
     2281 acgattttga ccttgacatg ctccccgggt aactaagtaa gaatccagac atgataagat
     2341 acattgatga gtttggacaa accacaacta gaatgcagtg aaaaaaatgc tttatttgtg
     2401 aaatttgtga tgctattgct ttatttgtaa ccattataag ctgcaataaa caagttaaca
     2461 acaacaattg cattcatttt atgtttcagg ttcaggggga ggtgtgggag gttttttaaa
     2521 gcaagtaaaa cctctacaaa tgtggtatgg ctgattatga tcctgcaagc ctcgtcgtcc
     2581 tggccggacc acgctatctg tgcaaggtcc ccggccccgg acgcgcgctc catgagcaga
     2641 gcgcccgccg ccgaggcgaa gactcgggcg gcgccctgcc cgtcccacca ggtcaacagg
     2701 cggtaaccgg cctcttcatc gggaatgcgc gcgaccttca gcatcgccgg catgtccccc
     2761 tggcggacgg gaagtatcca gctcgaccag gcttgtcgac tcgaagaacc cagctttctt
     2821 gtacaaagtt ggcattataa gaaagcattg cttatcaatt tgttgcaacg aacaggtcac
     2881 tatcagtcaa aataaaatca ttatttgcca tccagctgca gggcggccgc gatatcccct
     2941 atagtgagtc gtattacatg gtcatagctg tttcctggca gctctggccc gtgtctcaaa
     3001 atctctgatg ttacattgca caagataaaa atatatcatc atgaacaata aaactgtctg
     3061 cttacataaa cagtaataca aggggtgtta tgagccatat tcaacgggaa acgtcgaggc
     3121 cgcgattaaa ttccaacatg gatgctgatt tatatgggta taaatgggct cgcgataatg
     3181 tcgggcaatc aggtgcgaca atctatcgct tgtatgggaa gcccgatgcg ccagagttgt
     3241 ttctgaaaca tggcaaaggt agcgttgcca atgatgttac agatgagatg gtcagactaa
     3301 actggctgac ggaatttatg cctcttccga ccatcaagca ttttatccgt actcctgatg
     3361 atgcatggtt actcaccact gcgatccccg gaaaaacagc attccaggta ttagaagaat
     3421 atcctgattc aggtgaaaat attgttgatg cgctggcagt gttcctgcgc cggttgcatt
     3481 cgattcctgt ttgtaattgt ccttttaaca gcgatcgcgt atttcgtctc gctcaggcgc
     3541 aatcacgaat gaataacggt ttggttgatg cgagtgattt tgatgacgag cgtaatggct
     3601 ggcctgttga acaagtctgg aaagaaatgc ataaactttt gccattctca ccggattcag
     3661 tcgtcactca tggtgatttc tcacttgata accttatttt tgacgagggg aaattaatag
     3721 gttgtattga tgttggacga gtcggaatcg cagaccgata ccaggatctt gccatcctat
     3781 ggaactgcct cggtgagttt tctccttcat tacagaaacg gctttttcaa aaatatggta
     3841 ttgataatcc tgatatgaat aaattgcagt ttcatttgat gctcgatgag tttttctaat
     3901 cagaattggt taattggttg taacactggc agagcattac gctgacttga cgggacggcg
     3961 caagctcatg accaaaatcc cttaacgtga gttacgcgtc gttccactga gcgtcagacc
     4021 ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct
     4081 tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa
     4141 ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag
     4201 tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc
     4261 tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg
     4321 actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca
     4381 cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat
     4441 gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg
     4501 tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc
     4561 ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc
     4621 ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc
     4681 cttttgctca catgtt
//
