LOCUS       pDEST/LIfeAct-mCherry-N1                4831 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    pDEST/LIfeAct-mCherry-N1
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4831)
  AUTHORS   Smyth JW, Vogan JM, Buch PJ, Zhang SS, Fong TS, Hong TT, Shaw RM
  TITLE     Actin cytoskeleton rest stops regulate anterograde traffic of 
            connexin 43 vesicles to the plasma membrane.
  JOURNAL   Circ Res. 2012 Mar 30;110(7):978-89. Epub 2012 Feb 9.
  PUBMED    22328533
REFERENCE   2  (bases 1 to 4831)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            //gt12365.cn
FEATURES             Location/Qualifiers
     source          1..4831
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     enhancer        54..357
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        358..561
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     512..532
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    595..619
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             649..699
                     /codon_start=1
                     /product="peptide that binds filamentous actin (Riedl et 
                     al., 2008)"
                     /label=Lifeact
                     /note="first 17 amino acids of budding yeast Abp140"
                     /translation="MGVADLIKKFESISKEE"
     protein_bind    complement(713..737)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     regulatory      751..760
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             757..1467
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(902..920)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1170..1189
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    1612..1733
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1649..1668)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1703..1722
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1740..2195)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1877..1898)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2089..2108
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2222..2326
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2328..2685
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     primer_bind     complement(2351..2371)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2536..2671
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2598..2617
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2720..3514
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(2774..2793)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3384..3403
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3702..3721)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3746..3793
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4122..4710
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4611..4630
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 taatagtaat caattacggg gtcattagtt catagcccat atatggagtt ccgcgttaca
       61 taacttacgg taaatggccc gcctggctga ccgcccaacg acccccgccc attgacgtca
      121 ataatgacgt atgttcccat agtaacgcca atagggactt tccattgacg tcaatgggtg
      181 gagtatttac ggtaaactgc ccacttggca gtacatcaag tgtatcatat gccaagtacg
      241 ccccctattg acgtcaatga cggtaaatgg cccgcctggc attatgccca gtacatgacc
      301 ttatgggact ttcctacttg gcagtacatc tacgtattag tcatcgctat taccatggtg
      361 atgcggtttt ggcagtacat caatgggcgt ggatagcggt ttgactcacg gggatttcca
      421 agtctccacc ccattgacgt caatgggagt ttgttttggc accaaaatca acgggacttt
      481 ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg gcggtaggcg tgtacggtgg
      541 gaggtctata taagcagagc tggtttagtg aaccgtcaga tccgctagca tcaaacaagt
      601 ttgtacaaaa aagcaggctt taaaggaacc aattcagtcg actggatcat gggcgtggcc
      661 gacctgatca agaagttcga gagcatcagc aaggaggagt cgagatatct agacccagct
      721 ttcttgtaca aagtggttcg atgggatccc gccaccatgg tgagcaaggg cgaggaggat
      781 aacatggcca tcatcaagga gttcatgcgc ttcaaggtgc acatggaggg ctccgtgaac
      841 ggccacgagt tcgagatcga gggcgagggc gagggccgcc cctacgaggg cacccagacc
      901 gccaagctga aggtgaccaa gggtggcccc ctgcccttcg cctgggacat cctgtcccct
      961 cagttcatgt acggctccaa ggcctacgtg aagcaccccg ccgacatccc cgactacttg
     1021 aagctgtcct tccccgaggg cttcaagtgg gagcgcgtga tgaacttcga ggacggcggc
     1081 gtggtgaccg tgacccagga ctcctccctg caggacggcg agttcatcta caaggtgaag
     1141 ctgcgcggca ccaacttccc ctccgacggc cccgtaatgc agaagaagac catgggctgg
     1201 gaggcctcct ccgagcggat gtaccccgag gacggcgccc tgaagggcga gatcaagcag
     1261 aggctgaagc tgaaggacgg cggccactac gacgctgagg tcaagaccac ctacaaggcc
     1321 aagaagcccg tgcagctgcc cggcgcctac aacgtcaaca tcaagttgga catcacctcc
     1381 cacaacgagg actacaccat cgtggaacag tacgaacgcg ccgagggccg ccactccacc
     1441 ggcggcatgg acgagctgta caagtaagaa ttcgaagctg tacaagtaaa gcggccgcga
     1501 ctctagatca taatcagcca taccacattt gtagaggttt tacttgcttt aaaaaacctc
     1561 ccacacctcc ccctgaacct gaaacataaa atgaatgcaa ttgttgttgt taacttgttt
     1621 attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca
     1681 tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttaaggcgta
     1741 aattgtaagc gttaatattt tgttaaaatt cgcgttaaat ttttgttaaa tcagctcatt
     1801 ttttaaccaa taggccgaaa tcggcaaaat cccttataaa tcaaaagaat agaccgagat
     1861 agggttgagt gttgttccag tttggaacaa gagtccacta ttaaagaacg tggactccaa
     1921 cgtcaaaggg cgaaaaaccg tctatcaggg cgatggccca ctacgtgaac catcacccta
     1981 atcaagtttt ttggggtcga ggtgccgtaa agcactaaat cggaacccta aagggagccc
     2041 ccgatttaga gcttgacggg gaaagccggc gaacgtggcg agaaaggaag ggaagaaagc
     2101 gaaaggagcg ggcgctaggg cgctggcaag tgtagcggtc acgctgcgcg taaccaccac
     2161 acccgccgcg cttaatgcgc cgctacaggg cgcgtcaggt ggcacttttc ggggaaatgt
     2221 gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag
     2281 acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtcctg aggcggaaag
     2341 aaccagctgt ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc cccagcaggc
     2401 agaagtatgc aaagcatgca tctcaattag tcagcaacca ggtgtggaaa gtccccaggc
     2461 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac catagtcccg
     2521 cccctaactc cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat
     2581 ggctgactaa ttttttttat ttatgcagag gccgaggccg cctcggcctc tgagctattc
     2641 cagaagtagt gaggaggctt ttttggaggc ctaggctttt gcaaagatcg atcaagagac
     2701 aggatgagga tcgtttcgca tgattgaaca agatggattg cacgcaggtt ctccggccgc
     2761 ttgggtggag aggctattcg gctatgactg ggcacaacag acaatcggct gctctgatgc
     2821 cgccgtgttc cggctgtcag cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc
     2881 cggtgccctg aatgaactgc aagacgaggc agcgcggcta tcgtggctgg ccacgacggg
     2941 cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg ggaagggact ggctgctatt
     3001 gggcgaagtg ccggggcagg atctcctgtc atctcacctt gctcctgccg agaaagtatc
     3061 catcatggct gatgcaatgc ggcggctgca tacgcttgat ccggctacct gcccattcga
     3121 ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg atggaagccg gtcttgtcga
     3181 tcaggatgat ctggacgaag agcatcaggg gctcgcgcca gccgaactgt tcgccaggct
     3241 caaggcgagc atgcccgacg gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc
     3301 gaatatcatg gtggaaaatg gccgcttttc tggattcatc gactgtggcc ggctgggtgt
     3361 ggcggaccgc tatcaggaca tagcgttggc tacccgtgat attgctgaag agcttggcgg
     3421 cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat
     3481 cgccttctat cgccttcttg acgagttctt ctgagcggga ctctggggtt cgaaatgacc
     3541 gaccaagcga cgcccaacct gccatcacga gatttcgatt ccaccgccgc cttctatgaa
     3601 aggttgggct tcggaatcgt tttccgggac gccggctgga tgatcctcca gcgcggggat
     3661 ctcatgctgg agttcttcgc ccaccctagg gggaggctaa ctgaaacacg gaaggagaca
     3721 ataccggaag gaacccgcgc tatgacggca ataaaaagac agaataaaac gcacggtgtt
     3781 gggtcgtttg ttcataaacg cggggttcgg tcccagggct ggcactctgt cgatacccca
     3841 ccgagacccc attggggcca atacgcccgc gtttcttcct tttccccacc ccacccccca
     3901 agttcgggtg aaggcccagg gctcgcagcc aacgtcgggg cggcaggccc tgccatagcc
     3961 tcaggttact catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc
     4021 taggtgaaga tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc
     4081 cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg
     4141 cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg
     4201 gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca
     4261 aatactgttc ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg
     4321 cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg
     4381 tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga
     4441 acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac
     4501 ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat
     4561 ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc
     4621 tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga
     4681 tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc
     4741 ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg
     4801 gataaccgta ttaccgccat gcattagtta t
//