LOCUS       sequence-148011-        5721 bp    DNA     circular SYN 14-APR-2025
DEFINITION  mammalian expression of Mff fused to GFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5721)
  AUTHORS   Friedman JR, Lackner LL, West M, DiBenedetto JR, Nunnari J, Voeltz 
            GK
  TITLE     ER tubules mark sites of mitochondrial division.
  JOURNAL   Science. 2011 Oct 21;334(6054):358-62. doi: 10.1126/science.1207385.
            Epub 2011 Sep 1.
  PUBMED    21885730
REFERENCE   2  (bases 1 to 5721)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1126/science.1207385"; journalName: "Science"; date: 
            "2011-10-21- 21"; volume: "334"; issue: "6054"; pages: "358-62"
COMMENT     Sequence Label: GFP-Mff
FEATURES             Location/Qualifiers
     source          1..5721
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        68..371
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        372..575
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     526..546
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      614..623
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             620..1336
                     /codon_start=1
                     /product="Aequorea coerulescens GFP"
                     /label=AcGFP1
                     /note="mammalian codon-optimized"
                     /translation="MVSKGAELFTGIVPILIELNGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLSYGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFEDD
                     GNYKSRAEVKFEGDTLVNRIELTGTDFKEDGNILGNKMEYNYNAHNVYIMTDKAKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMIYF
                     GFVTAAAITHGMDELYK"
     polyA_signal    2516..2637
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2553..2572)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2607..2626
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2644..3099)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2781..2802)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2993..3012
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3126..3230
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3232..3589
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3232..3423
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3255..3275)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3440..3575
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3624..4418
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             3624..4418
                     /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(3678..3697)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4288..4307
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4606..4625)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4650..4697
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5026..5614
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5515..5534
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 catgcattag ttattaatag taatcaatta cggggtcatt agttcatagc ccatatatgg
       61 agttccgcgt tacataactt acggtaaatg gcccgcctgg ctgaccgccc aacgaccccc
      121 gcccattgac gtcaataatg acgtatgttc ccatagtaac gccaataggg actttccatt
      181 gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt ggcagtacat caagtgtatc
      241 atatgccaag tacgccccct attgacgtca atgacggtaa atggcccgcc tggcattatg
      301 cccagtacat gaccttatgg gactttccta cttggcagta catctacgta ttagtcatcg
      361 ctattaccat ggtgatgcgg ttttggcagt acatcaatgg gcgtggatag cggtttgact
      421 cacggggatt tccaagtctc caccccattg acgtcaatgg gagtttgttt tggcaccaaa
      481 atcaacggga ctttccaaaa tgtcgtaaca actccgcccc attgacgcaa atgggcggta
      541 ggcgtgtacg gtgggaggtc tatataagca gagctggttt agtgaaccgt cagatccgct
      601 agcgctaccg gtcgccacca tggtgagcaa gggcgccgag ctgttcaccg gcatcgtgcc
      661 catcctgatc gagctgaatg gcgatgtgaa tggccacaag ttcagcgtga gcggcgaggg
      721 cgagggcgat gccacctacg gcaagctgac cctgaagttc atctgcacca ccggcaagct
      781 gcctgtgccc tggcccaccc tggtgaccac cctgagctac ggcgtgcagt gcttctcacg
      841 ctaccccgat cacatgaagc agcacgactt cttcaagagc gccatgcctg agggctacat
      901 ccaggagcgc accatcttct tcgaggatga cggcaactac aagtcgcgcg ccgaggtgaa
      961 gttcgagggc gataccctgg tgaatcgcat cgagctgacc ggcaccgatt tcaaggagga
     1021 tggcaacatc ctgggcaata agatggagta caactacaac gcccacaatg tgtacatcat
     1081 gaccgacaag gccaagaatg gcatcaaggt gaacttcaag atccgccaca acatcgagga
     1141 tggcagcgtg cagctggccg accactacca gcagaatacc cccatcggcg atggccctgt
     1201 gctgctgccc gataaccact acctgtccac ccagagcgcc ctgtccaagg accccaacga
     1261 gaagcgcgat cacatgatct acttcggctt cgtgaccgcc gccgccatca cccacggcat
     1321 ggatgagctg tacaagtccg gactcagatc tcgagtgatg agtaaaggaa caagcagtga
     1381 cacatcacta ggaagggtga gcagggcagc atttccttct cccactgctg ctgagatggc
     1441 agaaattagt cgaattcagt acgaaatgga atatactgaa ggcattagtc agcgaatgag
     1501 ggtcccagaa aagttaaaag tagcaccgcc aaacgctgac ctggaacaag gattccaaga
     1561 aggagttcca aatgctagtg tgataatgca agttccggag aggattgttg tagcaggaaa
     1621 taatgaagat gtttcatttt caagaccagc agatcttgac cttattcagt caactccctt
     1681 taaacccctg gcactgaaaa caccacctcg tgtacttacg ctgagtgaaa gaccactaga
     1741 ttttctggat ttagaaagac ctcctacaac ccctcaaaat gaagaaatcc gagcagttgg
     1801 cagactaaaa agagagcggt ctatgagtga aaatgctgtt cgccaaaatg gacagctggt
     1861 cagaaatgat tctctgtggc acagatcaga ttctgcccca agaaataaaa tttcaaggtt
     1921 ccaggcaccg atttctgcac cggagtacac tgtgacacca tcgccacaac aggctcgggt
     1981 ctgtcctccc catatgttac ctgaagatgg agctaatctt tcctctgctc gtggcatttt
     2041 gtcgcttatc cagtcttcta ctcgtagggc ataccagcag atcttggatg tgctggatga
     2101 aaatcgcaga cctgtgttgc gtggtgggtc tgctgccgcc acttctaatc ctcatcatga
     2161 caacgtcagg tatggcattt caaatataga tacaaccatt gaaggaacgt cagatgacct
     2221 gactgttgta gatgcagctt cactaagacg acagataatc aaactaaata gacgtctaca
     2281 acttctggaa gaggagaaca aagaacgtgc taaaagagaa atggtcatgt attcaattac
     2341 tgtagctttc tggctgctta atagctggct ctggtttcgc cgctagggat ccaccggatc
     2401 tagataactg atcataatca gccataccac atttgtagag gttttacttg ctttaaaaaa
     2461 cctcccacac ctccccctga acctgaaaca taaaatgaat gcaattgttg ttgttaactt
     2521 gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa
     2581 agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttaacg
     2641 cgtaaattgt aagcgttaat attttgttaa aattcgcgtt aaatttttgt taaatcagct
     2701 cattttttaa ccaataggcc gaaatcggca aaatccctta taaatcaaaa gaatagaccg
     2761 agatagggtt gagtgttgtt ccagtttgga acaagagtcc actattaaag aacgtggact
     2821 ccaacgtcaa agggcgaaaa accgtctatc agggcgatgg cccactacgt gaaccatcac
     2881 cctaatcaag ttttttgggg tcgaggtgcc gtaaagcact aaatcggaac cctaaaggga
     2941 gcccccgatt tagagcttga cggggaaagc cggcgaacgt ggcgagaaag gaagggaaga
     3001 aagcgaaagg agcgggcgct agggcgctgg caagtgtagc ggtcacgctg cgcgtaacca
     3061 ccacacccgc cgcgcttaat gcgccgctac agggcgcgtc aggtggcact tttcggggaa
     3121 atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca
     3181 tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt cctgaggcgg
     3241 aaagaaccag ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag gctccccagc
     3301 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc
     3361 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccatagt
     3421 cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc
     3481 ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctcgg cctctgagct
     3541 attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaag atcgatcaag
     3601 agacaggatg aggatcgttt cgcatgattg aacaagatgg attgcacgca ggttctccgg
     3661 ccgcttgggt ggagaggcta ttcggctatg actgggcaca acagacaatc ggctgctctg
     3721 atgccgccgt gttccggctg tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc
     3781 tgtccggtgc cctgaatgaa ctgcaagacg aggcagcgcg gctatcgtgg ctggccacga
     3841 cgggcgttcc ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg gactggctgc
     3901 tattgggcga agtgccgggg caggatctcc tgtcatctca ccttgctcct gccgagaaag
     3961 tatccatcat ggctgatgca atgcggcggc tgcatacgct tgatccggct acctgcccat
     4021 tcgaccacca agcgaaacat cgcatcgagc gagcacgtac tcggatggaa gccggtcttg
     4081 tcgatcagga tgatctggac gaagagcatc aggggctcgc gccagccgaa ctgttcgcca
     4141 ggctcaaggc gagcatgccc gacggcgagg atctcgtcgt gacccatggc gatgcctgct
     4201 tgccgaatat catggtggaa aatggccgct tttctggatt catcgactgt ggccggctgg
     4261 gtgtggcgga ccgctatcag gacatagcgt tggctacccg tgatattgct gaagagcttg
     4321 gcggcgaatg ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc gattcgcagc
     4381 gcatcgcctt ctatcgcctt cttgacgagt tcttctgagc gggactctgg ggttcgaaat
     4441 gaccgaccaa gcgacgccca acctgccatc acgagatttc gattccaccg ccgccttcta
     4501 tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg
     4561 ggatctcatg ctggagttct tcgcccaccc tagggggagg ctaactgaaa cacggaagga
     4621 gacaataccg gaaggaaccc gcgctatgac ggcaataaaa agacagaata aaacgcacgg
     4681 tgttgggtcg tttgttcata aacgcggggt tcggtcccag ggctggcact ctgtcgatac
     4741 cccaccgaga ccccattggg gccaatacgc ccgcgtttct tccttttccc caccccaccc
     4801 cccaagttcg ggtgaaggcc cagggctcgc agccaacgtc ggggcggcag gccctgccat
     4861 agcctcaggt tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag
     4921 gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc
     4981 gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt
     5041 tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt
     5101 gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat
     5161 accaaatact gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc
     5221 accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa
     5281 gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg
     5341 ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag
     5401 atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag
     5461 gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa
     5521 cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt
     5581 gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg
     5641 gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc
     5701 tgtggataac cgtattaccg c
//
