LOCUS       Exported                4759 bp ds-DNA     circular SYN 21-DEC-2021
DEFINITION  Expresses a mitochondrial matrix-targeted photoactivable GFP that 
            enables detection and quantification of organelle fusion in living 
            cells..
ACCESSION   .
VERSION     .
KEYWORDS    mito-PAGFP
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4759)
  AUTHORS   Karbowski M, Arnoult D, Chen H, Chan DC, Smith CL, Youle RJ
  TITLE     Quantitation of mitochondrial dynamics by photolabeling of 
            individual organelles shows that mitochondrial fusion is blocked 
            during the Bax activation phase of apoptosis.
  JOURNAL   J Cell Biol. 2004 Feb 16. 164(4):493-9.
  PUBMED    14769861
REFERENCE   2  (bases 1 to 4759)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "J Cell 
            Biol. 2004 Feb 16. 164(4):493-9."
FEATURES             Location/Qualifiers
     source          1..4759
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     196..216
                     /label=pBABE 3
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        complement(241..345)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      372..827
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(459..478)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     669..690
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     polyA_signal    834..955
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(845..864)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     899..918
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(1081..1797)
                     /codon_start=1
                     /product="photoactivatable GFP variant (Patterson et al., 
                     2002)"
                     /label=PA-GFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFSYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSHQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1123..1144)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     1472..1491
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1731..1752
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      1794..1803
                     /regulatory_class="other"
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             complement(1831..1905)
                     /codon_start=1
                     /product="mitochondrial presequence of human cytochrome c 
                     oxidase subunit VIII (Rizutto et al., 1989)"
                     /label=COX8 presequence
                     /translation="MSVLTPLLLRGLTGSARRLPVPRAK"
     promoter        complement(1934..2137)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(1963..1983)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        2138..2441
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     rep_origin      complement(2616..3204)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2696..2715)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    3533..3580
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     primer_bind     3605..3624
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     CDS             complement(3812..4606)
                     /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(3923..3942)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     4533..4552
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(join(4641..4759,1..239))
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      join(4655..4759,1..31)
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(4709..4728)
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"