LOCUS       sequence-438069-        7568 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses mStayGold targeted to the mitochondria.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7568)
  TITLE     Janelia Integrative Imaging
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7568)
  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: 
            "Unpublished"
COMMENT     Sequence Label: mito-mStayGold_pTwistCMVBetaGlobWPRENeo
FEATURES             Location/Qualifiers
     source          1..7568
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             5..79
                     /codon_start=1
                     /product="mitochondrial presequence of human cytochrome c 
                     oxidase subunit VIII (Rizutto et al., 1989)"
                     /label=COX8 presequence
                     /translation="MSVLTPLLLRGLTGSARRLPVPRAK"
     CDS             107..781
                     /codon_start=1
                     /product="bright and highly photostable monomeric green 
                     fluorescent protein (Ando et al., 2023)"
                     /label=mStayGold(J)
                     /note="includes multiple mutations plus an N-terminal 
                     iinker sequence"
                     /translation="VSTGEELFTGVVPFKFQLKGTINGKSFTVEGEGEGNSHEGSHKGK
                     YVCTSGKLPMSWAALGTSFGYGMKYYTKYPSGLKNWFHEVMPEGFTYDRHIQYKGDGSI
                     HAKHQHFMKNGTYHNIVEFTGQDFKENSPVLTGDMDVSLPNEVQHIPIDDGVECTVTLQ
                     YPLLSDESKCVEAYQNTIIKPLHNQPAPDVPFHWIRKQYTQSKDDTEERDHIIQSETLE
                     AHL"
     misc_feature    848..1436
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(901..921)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(1319..1330)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    1489..1610
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1500..1519)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1554..1573
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      1725..1771
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      1801..1937
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    1982..2053
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     promoter        2098..2294
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      2145..2280
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2308..3102
                     /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
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(2362..2381)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2972..2991
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3113..3337
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      complement(3533..4121)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3613..3632)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        4867..4971
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4972..5040,5041..5832)
                     /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     complement(5190..5209)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      5845..5981
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    6126..6197
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(6202..6248)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    6266..6314
                     /note="synthetic polyadenylation signal"
     enhancer        6395..6698
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6700..6902
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     6853..6873
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     intron          7036..7511
                     /label=beta-globin intron
                     /note="internally truncated intron from human beta-globin"
ORIGIN
        1 caccatgtcc gtcctgacgc cgctgctgct gcggggcttg acaggctcgg cccggcggct
       61 cccagtgccg cgcgccaagg gagcatcggg agcctcagga gcatcggtgt ctacaggcga
      121 ggagctgttt accggcgtgg tgcccttcaa gttccagctg aagggcacca tcaacggcaa
      181 gagcttcacc gtggaagggg aaggtgaagg gaacagccac gagggcagcc acaaaggcaa
      241 gtacgtgtgc accagcggca aactgccaat gtcttgggcc gccctgggaa ctagcttcgg
      301 ctatggcatg aagtactaca ccaagtaccc cagcggcctg aagaactggt tccacgaggt
      361 gatgccggaa gggtttactt acgacagaca catccagtac aagggcgacg gcagcatcca
      421 cgccaagcac cagcacttca tgaagaacgg cacctaccac aacatcgtgg agttcaccgg
      481 ccaggacttc aaggagaaca gccccgtgct gaccggcgac atggacgtga gcctgcccaa
      541 cgaggtgcag cacatcccca tagatgacgg cgtggagtgc acagtgaccc tgcagtaccc
      601 tctgctgagc gacgagagca agtgcgtgga ggcctaccag aacaccatca tcaagcccct
      661 gcacaatcag ccagcccccg atgtgccatt ccactggatc agaaagcagt acacccagag
      721 caaggacgac accgaggaga gagaccacat catccagagc gagaccctgg aggcccacct
      781 gtaatctaga agttgtctcc tcctgcactg actgactgat acaatcgatt tctggatccg
      841 caggcctaat caacctctgg attacaaaat ttgtgaaaga ttgactggta ttcttaacta
      901 tgttgctcct tttacgctat gtggatacgc tgctttaatg cctttgtatc atgctattgc
      961 ttcccgtatg gctttcattt tctcctcctt gtataaatcc tggttgctgt ctctttatga
     1021 ggagttgtgg cccgttgtca ggcaacgtgg cgtggtgtgc actgtgtttg ctgacgcaac
     1081 ccccactggt tggggcattg ccaccacctg tcagctcctt tccgggactt tcgctttccc
     1141 cctccctatt gccacggcgg aactcatcgc cgcctgcctt gcccgctgct ggacaggggc
     1201 tcggctgttg ggcactgaca attccgtggt gttgtcgggg aagctgacgt cctttccatg
     1261 gctgctcgcc tgtgttgcca cctggattct gcgcgggacg tccttctgct acgtcccttc
     1321 ggccctcaat ccagcggacc ttccttcccg cggcctgctg ccggctctgc ggcctcttcc
     1381 gcgtcttcgc cttcgccctc agacgagtcg gatctccctt tgggccgcct ccccgcctgt
     1441 ctagcttgac tgactgagat acagcgtacc ttcagctcac agacatgata agatacattg
     1501 atgagtttgg acaaaccaca actagaatgc agtgaaaaaa atgctttatt tgtgaaattt
     1561 gtgatgctat tgctttattt gtaaccatta taagctgcaa taaacaagtt aacaacaaca
     1621 attgcattca ttttatgttt caggttcagg gggaggtgtg ggaggttttt taaagcaagt
     1681 aaaacctcta caaatgtggt attggcccat ctctatcggt atcgtagcat aaccccttgg
     1741 ggcctctaaa cgggtcttga ggggtttttt gtgcccctcg ggccggattg ctatctaccg
     1801 gcattggcgc agaaaaaaat gcctgatgcg acgctgcgcg tcttatactc ccacatatgc
     1861 cagattcagc aacggatacg gcttccccaa cttgcccact tccatacgtg tcctccttac
     1921 cagaaattta tccttaaggt cgtcagctat cctgcaggcg atctctcgat ttcgatcaag
     1981 acattccttt aatggtcttt tctggacacc actaggggtc agaagtagtt catcaaactt
     2041 tcttccctcc ctaatctcat tggttacctt gggctatcga aacttaatta agcgatctgc
     2101 atctcaatta gtcagcaacc atagtcccgc ccctaactcc gcccatcccg cccctaactc
     2161 cgcccagttc cgcccattct ccgccccatc gctgactaat tttttttatt tatgcagagg
     2221 ccgaggccgc ctcggcctct gagctattcc agaagtagtg aggaggcttt tttggaggcc
     2281 taggcttttg caaaggaggt agccaacatg attgaacaag atggattgca cgcaggttct
     2341 cccgccgctt gggtggagag gctattcggc tatgactggg cacaacagac aatcggctgc
     2401 tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc cggttctttt tgtcaagacc
     2461 gacctgtccg gtgccctgaa tgaactccag gacgaggcag cgcggctatc gtggctggcc
     2521 acgacgggcg ttccttgcgc agctgtgctc gacgttgtca ctgaagcggg aagggactgg
     2581 ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat ctcaccttgc tcctgccgag
     2641 aaagtatcca tcatggctga tgcaatgcgg cggctgcata cgcttgatcc ggctacctgc
     2701 ccattcgacc accaagcgaa acatcgcatc gagcgagcac gtactcggat ggaagccggt
     2761 cttgtcgatc aggatgatct ggacgaagag catcaggggc tcgcgccagc cgaactgttc
     2821 gccaggctca aggcgcggat gcccgacggc gaggatctcg tcgtgaccca cggcgatgcc
     2881 tgcttgccga atatcatggt ggaaaatggc cgcttttctg gattcatcga ctgtggccgg
     2941 ctgggtgtgg cggaccgcta tcaggacata gcgttggcta cccgtgatat tgctgaagag
     3001 cttggcggcg aatgggctga ccgcttcctc gtgctttacg gtatcgccgc tcccgattcg
     3061 cagcgcatcg ccttctatcg ccttcttgac gagttcttct agtatgtaag ccctgtgcct
     3121 tctagttgcc agccatctgt tgtttgcccc tcccccgtgc cttccttgac cctggaaggt
     3181 gccactccca ctgtcctttc ctaataaaat gaggaaattg catcgcattg tctgagtagg
     3241 tgtcattcta ttctgggggg tggggtgggg caggacagca agggggagga ttgggaagac
     3301 aatagcaggc atgctgggga tgcggtgggc tctatggtta attaaccagt caagtcagct
     3361 acttggcgag atcgacttgt ctgggtttcg actacgctca gaattgcgtc agtcaagttc
     3421 gatctggtcc ttgctattgc acccgttctc cgattacgag tttcatttaa atcatgtgag
     3481 caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata
     3541 ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc
     3601 cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg
     3661 ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc
     3721 tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg
     3781 gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc
     3841 ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga
     3901 ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg
     3961 gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa
     4021 aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg
     4081 tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt
     4141 ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat
     4201 tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct
     4261 aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta
     4321 tctcagcgat ctgtctattt cgttcatcca tagttgcatt taaatttccg aactctccaa
     4381 ggccctcgtc ggaaaatctt caaacctttc gtccgatcca tcttgcaggc tacctctcga
     4441 acgaactatc gcaagtctct tggccggcct tgcgccttgg ctattgcttg gcagcgccta
     4501 tcgccaggta ttactccaat cccgaatatc cgagatcggg atcacccgag agaagttcaa
     4561 cctacatcct caatcccgat ctatccgaga tccgaggaat atcgaaatcg gggcgcgcct
     4621 ggtgtaccga gaacgatcct ctcagtgcga gtctcgacga tccatatcgt tgcttggcag
     4681 tcagccagtc ggaatccagc ttgggaccca ggaagtccaa tcgtcagata ttgtactcaa
     4741 gcctggtcac ggcagcgtac cgatctgttt aaacctagat attgatagtc tgatcggtca
     4801 acgtataatc gagtcctagc ttttgcaaac atctatcaag agacaggatc agcaggaggc
     4861 tttcgccgcg gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc
     4921 atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag tatgagtatt
     4981 caacatttcc gtgtcgccct tattcccttt tttgcggcat tttgccttcc tgtttttgct
     5041 cacccagaaa cgctggtgaa agtaaaagat gctgaagatc agttgggtgc gcgagtgggt
     5101 tacatcgaac tggatctcaa cagcggtaag atccttgaga gttttcgccc cgaagaacgc
     5161 tttccaatga tgagcacttt taaagttctg ctatgtggcg cggtattatc ccgtattgac
     5221 gccgggcaag agcaactcgg tcgccgcata cactattctc agaatgactt ggttgagtat
     5281 tcaccagtca cagaaaagca tcttacggat ggcatgacag taagagaatt atgcagtgct
     5341 gccataacca tgagtgataa cactgcggcc aacttacttc tgacaacgat tggaggaccg
     5401 aaggagctaa ccgctttttt gcacaacatg ggggatcatg taactcgcct tgatcgttgg
     5461 gaaccggagc tgaatgaagc cataccaaac gacgagcgtg acaccacgat gcctgtagca
     5521 atggcaacaa ccttgcgtaa actattaact ggcgaactac ttactctagc ttcccggcaa
     5581 cagttgatag actggatgga ggcggataaa gttgcaggac cacttctgcg ctcggccctt
     5641 ccggctggct ggtttattgc tgataaatct ggagccggtg agcgtgggtc tcgcggtatc
     5701 attgcagcac tggggccaga tggtaagccc tcccgtatcg tagttatcta cacgacgggg
     5761 agtcaggcaa ctatggatga acgaaataga cagatcgctg agataggtgc ctcactgatt
     5821 aagcattggt aaccgattct aggtgcattg gcgcagaaaa aaatgcctga tgcgacgctg
     5881 cgcgtcttat actcccacat atgccagatt cagcaacgga tacggcttcc ccaacttgcc
     5941 cacttccata cgtgtcctcc ttaccagaaa tttatcctta agatcgttta aactcgactc
     6001 tggctctatc gaatctccgt cgtttcgagc ttacgcgaac agccgtggcg ctcatttgct
     6061 cgtcgggcat cgaatctcgt cagctatcgt cagcttacct ttttggcagc gatcgcggct
     6121 cccgacatct tggaccatta gctccacagg tatcttcttc cctctagtgg tcataacagc
     6181 agcttcagct acctctcaat tcaaaaaacc cctcaagacc cgtttagagg ccccaagggg
     6241 ttatgctatc aatcgttgcg ttacacacac aaaaaaccaa cacacatcca tcttcgatgg
     6301 atagcgattt tattatctaa ctgctgatcg agtgtagcca gatctagtaa tcaattacgg
     6361 ggtcattagt tcatagccca tatatggagt tccgcgttac ataacttacg gtaaatggcc
     6421 cgcctggctg accgcccaac gacccccgcc cattgacgtc aataatgacg tatgttccca
     6481 tagtaacgcc aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg
     6541 cccacttggc agtacatcaa gtgtatcata tgccaagtac gccccctatt gacgtcaatg
     6601 acggtaaatg gcccgcctgg cattatgccc agtacatgac cttatgggac tttcctactt
     6661 ggcagtacat ctacgtatta gtcatcgcta ttaccatgct gatgcggttt tggcagtaca
     6721 tcaatgggcg tggatagcgg tttgactcac ggggatttcc aagtctccac cccattgacg
     6781 tcaatgggag tttgttttgg caccaaaatc aacgggactt tccaaaatgt cgtaacaact
     6841 ccgccccatt gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat ataagcagag
     6901 ctggtttagt gaaccgtcag atcgcctgga gacgccatcc acgctgtttt gacctccata
     6961 gaagacaccg ggaccgatcc agcctcccct cgaagcttac atgtggtacc gagctcggat
     7021 cctgagaact tcagggtgag tctatgggac ccttgatgtt ttctttcccc ttcttttcta
     7081 tggttaagtt catgtcatag gaaggggaga agtaacaggg tacacatatt gaccaaatca
     7141 gggtaatttt gcatttgtaa ttttaaaaaa tgctttcttc ttttaatata cttttttgtt
     7201 tatcttattt ctaatacttt ccctaatctc tttctttcag ggcaataatg atacaatgta
     7261 tcatgcctct ttgcaccatt ctaaagaata acagtgataa tttctgggtt aaggcaatag
     7321 caatatttct gcatataaat atttctgcat ataaattgta actgatgtaa gaggtttcat
     7381 attgctaata gcagctacaa tccagctacc attctgcttt tattttatgg ttgggataag
     7441 gctggattat tctgagtcca agctaggccc ttttgctaat catgttcata cctcttatct
     7501 tcctcccaca gctcctgggc aacgtgctgg tctgtgtgct ggcccatcac tttggcaaag
     7561 gcggccgc
//
