LOCUS       sequence-477204-        8813 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Cytosolic expression of mCherry.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8813)
  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 8813)
  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 mCherry-P2A-NeoR
FEATURES             Location/Qualifiers
     source          1..8813
                     /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"
     regulatory      3864..3873
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3870..4577
                     /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(4015..4033)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     4283..4302
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             4602..4658
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             4653..5453
                     /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="GPMIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQG
                     RPVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDL
                     LSSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEE
                     HQGLAPAELFARLKARMPDGDDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQD
                     IALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4713..4732)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5323..5342
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     misc_feature    5479..6067
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(5532..5552)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5950..5961)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(6070..6086)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6071..6087)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             6592..6772
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     primer_bind     complement(6798..6815)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6804..6915
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      complement(7061..7649)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(7141..7160)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7820..8611,8612..8680))
                     /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     8443..8462
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8681..8785)
                     /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 aaggccacca tggtgagcaa gggcgaggag gataacatgg
     3901 ccatcatcaa ggagttcatg cgcttcaagg tgcacatgga gggctccgtg aacggccacg
     3961 agttcgagat cgagggcgag ggcgagggcc gcccctacga gggcacccag accgccaagc
     4021 tgaaggtgac caagggtggc cccctgccct tcgcctggga catcctgtcc cctcagttca
     4081 tgtacggctc caaggcctac gtgaagcacc ccgccgacat ccccgactac ttgaagctgt
     4141 ccttccccga gggcttcaag tgggagcgcg tgatgaactt cgaggacggc ggcgtggtga
     4201 ccgtgaccca ggactcctcc ctgcaggacg gcgagttcat ctacaaggtg aagctgcgcg
     4261 gcaccaactt cccctccgac ggccccgtaa tgcagaagaa gaccatgggc tgggaggcct
     4321 cctccgagcg gatgtacccc gaggacggcg ccctgaaggg cgagatcaag cagaggctga
     4381 agctgaagga cggcggccac tacgacgctg aggtcaagac cacctacaag gccaagaagc
     4441 ccgtgcagct gcccggcgcc tacaacgtca acatcaagtt ggacatcacc tcccacaacg
     4501 aggactacac catcgtggaa cagtacgaac gcgccgaggg ccgccactcc accggcggca
     4561 tggacgagct gtacaaggat ctgcagtcta gaggatctgg tgccacgaat ttctcactac
     4621 tcaagcaggc cggtgatgtc gaggaaaacc ctggtcctat gattgaacaa gatggattgc
     4681 acgcaggttc tccggccgct tgggtggaga ggctattcgg ctatgactgg gcacaacaga
     4741 caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt
     4801 ttgtcaagac cgacctgtcc ggtgccctga atgaactgca ggacgaggca gcgcggctat
     4861 cgtggctggc cacgacgggc gttccttgcg cagctgtgct cgacgttgtc actgaagcgg
     4921 gaagggactg gctgctattg ggcgaagtgc cggggcagga tctcctgtca tctcaccttg
     4981 ctcctgccga gaaagtatcc atcatggctg atgcaatgcg gcggctgcat acgcttgatc
     5041 cggctacctg cccattcgac caccaagcga aacatcgcat cgagcgagca cgtactcgga
     5101 tggaagccgg tcttgtcgat caggatgatc tggacgaaga gcatcagggg ctcgcgccag
     5161 ccgaactgtt cgccaggctc aaggcgcgca tgcccgacgg cgatgatctc gtcgtgaccc
     5221 atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct ggattcatcg
     5281 actgtggccg gctgggtgtg gcggaccgct atcaggacat agcgttggct acccgtgata
     5341 ttgctgaaga gcttggcggc gaatgggctg accgcttcct cgtgctttac ggtatcgccg
     5401 ctcccgattc gcagcgcatc gccttctatc gccttcttga cgagttcttc tgacgaattc
     5461 gatatcaagc ttatcgataa tcaacctctg gattacaaaa tttgtgaaag attgactggt
     5521 attcttaact atgttgctcc ttttacgcta tgtggatacg ctgctttaat gcctttgtat
     5581 catgctattg cttcccgtat ggctttcatt ttctcctcct tgtataaatc ctggttgctg
     5641 tctctttatg aggagttgtg gcccgttgtc aggcaacgtg gcgtggcgtg cactgtgttt
     5701 gctgacgcaa cccccactgg ttggggcatt gccaccacct gtcagctcct ttccgggact
     5761 ttcgctttcc ccctccctat tgccacggcg gaactcatcg ccgcctgcct tgcccgctgc
     5821 tggacagggg ctcggctgtt gggcactgac aattccgtgg tgttgtcggg gaaatcatcg
     5881 tcctttcctt ggctgctcgc ctgtgttgcc acctggattc tgcgcgggac gtccttctgc
     5941 tacgtccctt cggccctcaa tccagcggac cttccttccc gcggcctgct gccggctctg
     6001 cggcctcttc cgcgtcttcg ccttcgccct cagacgagtc ggatctccct ttgggccgcc
     6061 tccccgcatc gataccgtcg acctcgagac ctagaaaaac atggagcaat cacaagtagc
     6121 aatacagcag ctaccaatgc tgattgtgcc tggctagaag cacaagagga ggaggaggtg
     6181 ggttttccag tcacacctca ggtaccttta agaccaatga cttacaaggc agctgtagat
     6241 cttagccact ttttaaaaga aaagggggga ctggaagggc taattcactc ccaacgaaga
     6301 caagatatcc ttgatctgtg gatctaccac acacaaggct acttccctga ttggcagaac
     6361 tacacaccag ggccagggat cagatatcca ctgacctttg gatggtgcta caagctagta
     6421 ccagttgagc aagagaaggt agaagaagcc aatgaaggag agaacacccg cttgttacac
     6481 cctgtgagcc tgcatgggat ggatgacccg gagagagaag tattagagtg gaggtttgac
     6541 agccgcctag catttcatca catggcccga gagctgcatc cggactgtac tgggtctctc
     6601 tggttagacc agatctgagc ctgggagctc tctggctaac tagggaaccc actgcttaag
     6661 cctcaataaa gcttgccttg agtgcttcaa gtagtgtgtg cccgtctgtt gtgtgactct
     6721 ggtaactaga gatccctcag acccttttag tcagtgtgga aaatctctag cagggcccgt
     6781 ttaaacccgc tgatcagcct cgactgtgcc ttctagttgc cagccatctg ttgtttgccc
     6841 ctcccccgtg ccttccttga ccctggaagg tgccactccc actgtccttt cctaataaaa
     6901 tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct attctggggg gtggggtggg
     6961 gcaggacagc aagggggagg attgggaaga caatagcagg catgtgagca aaaggccagc
     7021 aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc
     7081 ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat
     7141 aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc
     7201 cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct
     7261 cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg
     7321 aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc
     7381 cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga
     7441 ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa
     7501 gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta
     7561 gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc
     7621 agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg
     7681 acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga
     7741 tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg
     7801 agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct
     7861 gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg
     7921 agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc
     7981 cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa
     8041 ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc
     8101 cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt
     8161 cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc
     8221 ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt
     8281 tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc
     8341 catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt
     8401 gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata
     8461 gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga
     8521 tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag
     8581 catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa
     8641 aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt
     8701 attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga
     8761 aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct gac
//
