LOCUS       sequence-477009-        8072 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression and mitochondrial localization of mEmerald.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8072)
  AUTHORS   Goldman, C., Kareva, T., Sarrafha, L., Schuldt, B.R., Sahasrabudhe, 
            A., Ahfeldt, T.,, Blanchard, J.W
  TITLE     Genetically encoded and modular subcellular organelle probes reveal 
            dysfunction in lysosomes and mitochondria driven by PRKN knockout
  JOURNAL   iScience 28 (2025)
REFERENCE   2  (bases 1 to 8072)
  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: "iScience 28
            (2025)"
COMMENT     Sequence Label: pFUW COX8ASig-mEmerald
FEATURES             Location/Qualifiers
     source          1..8072
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(47..66)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        238..617
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        619..817
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     772..792
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             835..1015
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1062..1187
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1680..1913
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2098..2142
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     misc_feature    2440..2557
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2622..3833
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     3670..3689
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             3879..3953
                     /codon_start=1
                     /product="mitochondrial presequence of human cytochrome c 
                     oxidase subunit VIII (Rizutto et al., 1989)"
                     /label=COX8 presequence
                     /translation="MSVLTPLLLRGLTGSARRLPVPRAK"
     regulatory      3981..3990
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(3987..3989,3990..3992,3993..4706)
                     /codon_start=1
                     /product="brightly fluorescent monomeric variant of GFP"
                     /label=mEmerald
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHKVYITADKQKNGIK
                     VNFKTRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(4032..4053)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4293..4312)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4640..4661
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    4738..5326
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4791..4811)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5209..5220)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(5329..5345)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5330..5346)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             5851..6031
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     primer_bind     complement(6057..6074)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6063..6174
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      complement(6320..6908)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6400..6419)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7079..7870,7871..7939))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     7702..7721
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7940..8044)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gtcgacggat cgggagatct cccgatcccc tatggtgcac tctcagtaca atctgctctg
       61 atgccgcata gttaagccag tatctgctcc ctgcttgtgt gttggaggtc gctgagtagt
      121 gcgcgagcaa aatttaagct acaacaaggc aaggcttgac cgacaattgc atgaagaatc
      181 tgcttagggt taggcgtttt gcgctgcttc gcgatgtacg ggccagatat acgcgttgac
      241 attgattatt gactagttat taatagtaat caattacggg gtcattagtt catagcccat
      301 atatggagtt ccgcgttaca taacttacgg taaatggccc gcctggctga ccgcccaacg
      361 acccccgccc attgacgtca ataatgacgt atgttcccat agtaacgcca atagggactt
      421 tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag
      481 tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc
      541 attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag
      601 tcatcgctat taccatggtg atgcggtttt ggcagtacat caatgggcgt ggatagcggt
      661 ttgactcacg gggatttcca agtctccacc ccattgacgt caatgggagt ttgttttggc
      721 accaaaatca acgggacttt ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg
      781 gcggtaggcg tgtacggtgg gaggtctata taagcagcgc gttttgcctg tactgggtct
      841 ctctggttag accagatctg agcctgggag ctctctggct aactagggaa cccactgctt
      901 aagcctcaat aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac
      961 tctggtaact agagatccct cagacccttt tagtcagtgt ggaaaatctc tagcagtggc
     1021 gcccgaacag ggacttgaaa gcgaaaggga aaccagagga gctctctcga cgcaggactc
     1081 ggcttgctga agcgcgcacg gcaagaggcg aggggcggcg actggtgagt acgccaaaaa
     1141 ttttgactag cggaggctag aaggagagag atgggtgcga gagcgtcagt attaagcggg
     1201 ggagaattag atcgcgatgg gaaaaaattc ggttaaggcc agggggaaag aaaaaatata
     1261 aattaaaaca tatagtatgg gcaagcaggg agctagaacg attcgcagtt aatcctggcc
     1321 tgttagaaac atcagaaggc tgtagacaaa tactgggaca gctacaacca tcccttcaga
     1381 caggatcaga agaacttaga tcattatata atacagtagc aaccctctat tgtgtgcatc
     1441 aaaggataga gataaaagac accaaggaag ctttagacaa gatagaggaa gagcaaaaca
     1501 aaagtaagac caccgcacag caagcggccg ctgatcttca gacctggagg aggagatatg
     1561 agggacaatt ggagaagtga attatataaa tataaagtag taaaaattga accattagga
     1621 gtagcaccca ccaaggcaaa gagaagagtg gtgcagagag aaaaaagagc agtgggaata
     1681 ggagctttgt tccttgggtt cttgggagca gcaggaagca ctatgggcgc agcgtcaatg
     1741 acgctgacgg tacaggccag acaattattg tctggtatag tgcagcagca gaacaatttg
     1801 ctgagggcta ttgaggcgca acagcatctg ttgcaactca cagtctgggg catcaagcag
     1861 ctccaggcaa gaatcctggc tgtggaaaga tacctaaagg atcaacagct cctggggatt
     1921 tggggttgct ctggaaaact catttgcacc actgctgtgc cttggaatgc tagttggagt
     1981 aataaatctc tggaacagat ttggaatcac acgacctgga tggagtggga cagagaaatt
     2041 aacaattaca caagcttaat acactcctta attgaagaat cgcaaaacca gcaagaaaag
     2101 aatgaacaag aattattgga attagataaa tgggcaagtt tgtggaattg gtttaacata
     2161 acaaattggc tgtggtatat aaaattattc ataatgatag taggaggctt ggtaggttta
     2221 agaatagttt ttgctgtact ttctatagtg aatagagtta ggcagggata ttcaccatta
     2281 tcgtttcaga cccacctccc aaccccgagg ggacccgaca ggcccgaagg aatagaagaa
     2341 gaaggtggag agagagacag agacagatcc attcgattag tgaacggatc ggcactgcgt
     2401 gcgccaattc tgcagacaaa tggcagtatt catccacaat tttaaaagaa aaggggggat
     2461 tggggggtac agtgcagggg aaagaatagt agacataata gcaacagaca tacaaactaa
     2521 agaattacaa aaacaaatta caaaaattca aaattttcgg gtttattaca gggacagcag
     2581 agatccagtt tggttaatta acccgtgtcg gctccagatc tggcctccgc gccgggtttt
     2641 ggcgcctccc gcgggcgccc ccctcctcac ggcgagcgct gccacgtcag acgaagggcg
     2701 cagcgagcgt cctgatcctt ccgcccggac gctcaggaca gcggcccgct gctcataaga
     2761 ctcggcctta gaaccccagt atcagcagaa ggacatttta ggacgggact tgggtgactc
     2821 tagggcactg gttttctttc cagagagcgg aacaggcgag gaaaagtagt cccttctcgg
     2881 cgattctgcg gagggatctc cgtggggcgg tgaacgccga tgattatata aggacgcgcc
     2941 gggtgtggca cagctagttc cgtcgcagcc gggatttggg tcgcggttct tgtttgtgga
     3001 tcgctgtgat cgtcacttgg tgagtagcgg gctgctgggc tggccggggc tttcgtggcc
     3061 gccgggccgc tcggtgggac ggaagcgtgt ggagagaccg ccaagggctg tagtctgggt
     3121 ccgcgagcaa ggttgccctg aactgggggt tggggggagc gcagcaaaat ggcggctgtt
     3181 cccgagtctt gaatggaaga cgcttgtgag gcgggctgtg aggtcgttga aacaaggtgg
     3241 ggggcatggt gggcggcaag aacccaaggt cttgaggcct tcgctaatgc gggaaagctc
     3301 ttattcgggt gagatgggct ggggcaccat ctggggaccc tgacgtgaag tttgtcactg
     3361 actggagaac tcggtttgtc gtctgttgcg ggggcggcag ttatggcggt gccgttgggc
     3421 agtgcacccg tacctttggg agcgcgcgcc ctcgtcgtgt cgtgacgtca cccgttctgt
     3481 tggcttataa tgcagggtgg ggccacctgc cggtaggtgt gcggtaggct tttctccgtc
     3541 gcaggacgca gggttcgggc ctagggtagg ctctcctgaa tcgacaggcg ccggacctct
     3601 ggtgagggga gggataagtg aggcgtcagt ttctttggtc ggttttatgt acctatcttc
     3661 ttaagtagct gaagctccgg ttttgaacta tgcgctcggg gttggcgagt gtgttttgtg
     3721 aagtttttta ggcacctttt gaaatgtaat catttgggtc aatatgtaat tttcagtgtt
     3781 agactagtaa attgtccgct aaattctggc cgtttttggc ttttttgtta gacgaagctt
     3841 gggcccggga attaattatt aaggtcagat ccgctagcat gtccgtcctg acgccgctgc
     3901 tgctgcgggg cttgacaggc tcggcccggc ggctcccagt gccgcgcgcc aagatccatt
     3961 cgttggggga tccaccggtc gccaccatgg tgagcaaggg cgaggagctg ttcaccgggg
     4021 tggtgcccat cctggtcgag ctggacggcg acgtaaacgg ccacaagttc agcgtgtccg
     4081 gcgagggcga gggcgatgcc acctacggca agctgaccct gaagttcatc tgcaccaccg
     4141 gcaagctgcc cgtgccctgg cccaccctcg tgaccacctt gacctacggc gtgcagtgct
     4201 tcgcccgcta ccccgaccac atgaagcagc acgacttctt caagtccgcc atgcccgaag
     4261 gctacgtcca ggagcgcacc atcttcttca aggacgacgg caactacaag acccgcgccg
     4321 aggtgaagtt cgagggcgac accctggtga accgcatcga gctgaagggc atcgacttca
     4381 aggaggacgg caacatcctg gggcacaagc tggagtacaa ctacaacagc cacaaggtct
     4441 atatcaccgc cgacaagcag aagaacggca tcaaggtgaa cttcaagacc cgccacaaca
     4501 tcgaggacgg cagcgtgcag ctcgccgacc actaccagca gaacaccccc atcggcgacg
     4561 gccccgtgct gctgcccgac aaccactacc tgagcaccca gtccaagctg agcaaagacc
     4621 ccaacgagaa gcgcgatcac atggtcctgc tggagttcgt gaccgccgcc gggatcactc
     4681 tcggcatgga cgagctgtac aagtaacgga tccgaattcg atatcaagct tatcgataat
     4741 caacctctgg attacaaaat ttgtgaaaga ttgactggta ttcttaacta tgttgctcct
     4801 tttacgctat gtggatacgc tgctttaatg cctttgtatc atgctattgc ttcccgtatg
     4861 gctttcattt tctcctcctt gtataaatcc tggttgctgt ctctttatga ggagttgtgg
     4921 cccgttgtca ggcaacgtgg cgtggtgtgc actgtgtttg ctgacgcaac ccccactggt
     4981 tggggcattg ccaccacctg tcagctcctt tccgggactt tcgctttccc cctccctatt
     5041 gccacggcgg aactcatcgc cgcctgcctt gcccgctgct ggacaggggc tcggctgttg
     5101 ggcactgaca attccgtggt gttgtcgggg aaatcatcgt cctttccttg gctgctcgcc
     5161 tgtgttgcca cctggattct gcgcgggacg tccttctgct acgtcccttc ggccctcaat
     5221 ccagcggacc ttccttcccg cggcctgctg ccggctctgc ggcctcttcc gcgtcttcgc
     5281 cttcgccctc agacgagtcg gatctccctt tgggccgcct ccccgcatcg ataccgtcga
     5341 cctcgagacc tagaaaaaca tggagcaatc acaagtagca atacagcagc taccaatgct
     5401 gattgtgcct ggctagaagc acaagaggag gaggaggtgg gttttccagt cacacctcag
     5461 gtacctttaa gaccaatgac ttacaaggca gctgtagatc ttagccactt tttaaaagaa
     5521 aaggggggac tggaagggct aattcactcc caacgaagac aagatatcct tgatctgtgg
     5581 atctaccaca cacaaggcta cttccctgat tggcagaact acacaccagg gccagggatc
     5641 agatatccac tgacctttgg atggtgctac aagctagtac cagttgagca agagaaggta
     5701 gaagaagcca atgaaggaga gaacacccgc ttgttacacc ctgtgagcct gcatgggatg
     5761 gatgacccgg agagagaagt attagagtgg aggtttgaca gccgcctagc atttcatcac
     5821 atggcccgag agctgcatcc ggactgtact gggtctctct ggttagacca gatctgagcc
     5881 tgggagctct ctggctaact agggaaccca ctgcttaagc ctcaataaag cttgccttga
     5941 gtgcttcaag tagtgtgtgc ccgtctgttg tgtgactctg gtaactagag atccctcaga
     6001 cccttttagt cagtgtggaa aatctctagc agggcccgtt taaacccgct gatcagcctc
     6061 gactgtgcct tctagttgcc agccatctgt tgtttgcccc tcccccgtgc cttccttgac
     6121 cctggaaggt gccactccca ctgtcctttc ctaataaaat gaggaaattg catcgcattg
     6181 tctgagtagg tgtcattcta ttctgggggg tggggtgggg caggacagca agggggagga
     6241 ttgggaagac aatagcaggc atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa
     6301 aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc
     6361 gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc
     6421 ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg
     6481 cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt
     6541 cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc
     6601 gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc
     6661 cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag
     6721 agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg
     6781 ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa
     6841 ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag
     6901 gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact
     6961 cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa
     7021 attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt
     7081 accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag
     7141 ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca
     7201 gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc
     7261 agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt
     7321 ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg
     7381 ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca
     7441 gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg
     7501 ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca
     7561 tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg
     7621 tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct
     7681 cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca
     7741 tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca
     7801 gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg
     7861 tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac
     7921 ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt
     7981 attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc
     8041 cgcgcacatt tccccgaaaa gtgccacctg ac
//
