LOCUS       sequence-216066-        6055 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of mitochondrially targeted, mVenus labelled 
            BoLD.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6055)
  AUTHORS   Reis JM, Xu X, McDonald S, Woloschuk RM, Jaikaran ASI, Vizeacoumar 
            FS, Woolley GA, Uppalapati M
  TITLE     Discovering Selective Binders for Photoswitchable Proteins Using 
            Phage Display.
  JOURNAL   ACS Synth Biol. 2018 Oct 19;7(10):2355-2364. doi: 
            10.1021/acssynbio.8b00123. Epub 2018 Sep 27.
  PUBMED    30203962
REFERENCE   2  (bases 1 to 6055)
  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"; doi: 
            "10.1021/acssynbio.8b00123"; journalName: "ACS Synth Biol"; date: 
            "2018-10-19- 19"; volume: "7"; issue: "10"; pages: "2355-2364"
COMMENT     Sequence Label: pTriEX Tom20 mVenus BoLD
FEATURES             Location/Qualifiers
     source          1..6055
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_recomb     110..937
                     /label=baculovirus recombination region (lef2/ORF603)
                     /note="contains ORF603 and part of lef2"
     CDS             110..234
                     /codon_start=3
                     /product="baculovirus late expression factor 2"
                     /label=lef2
                     /translation="HLLIKSQDVYKPPNCQKMKTVDKLCPFAGNCKGLNPICNY"
     CDS             complement(272..877)
                     /codon_start=1
                     /product="baculovirus ORF603 protein"
                     /label=ORF603
                     /translation="MAVIFNNKQLLADNSIEKGGELFLFNGSYNILESYVNPVLLKNGV
                     IELEEAAYYAGNILYKTDDPKFIDYINLIIKATHSEELPENSTVVNYRKTMRSGTIHPI
                     KKDIYIYDNKKFTLYDRYIYGYDNNYVNFYEEKNEKEKEYEEEDDKASSLCENKIILSQ
                     INCESFENDFKYYLSDYNYAFSIIDNTTNVLVAFGLYR"
     enhancer        1005..1308
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1310..1512
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1463..1483
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     1715..1737
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1799..1818
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     primer_bind     1860..1879
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1860..1878
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1879..1903
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        1919..2028
                     /label=p10 promoter
                     /note="baculovirus promoter for expression in insect cells"
     CDS             join(2154..2156,2157..2867)
                     /codon_start=1
                     /product="photostable derivative of mVenus (Lee et al., 
                     2020)"
                     /label=mGold
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FISTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIKA
                     NFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSKLSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(2196..2217)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2457..2476)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2804..2825
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             3084..3101
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             3108..3128
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             3135..3158
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             3198..3230
                     /codon_start=1
                     /product="HSV (herpes simplex virus) epitope tag"
                     /label=HSV tag
                     /translation="QPELAPEDPED"
     CDS             3237..3260
                     /codon_start=1
                     /product="8xHis affinity tag"
                     /label=8xHis
                     /translation="HHHHHHHH"
     primer_bind     complement(3350..3369)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    3415..3470
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(3469..3488)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     terminator      3558..3605
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     misc_recomb     3618..4323
                     /label=baculovirus recombination region (ORF1629)
                     /note="contains part of ORF1629"
     3'UTR           3706..4078
                     /label=polyhedrin 3' UTR
                     /note="3' UTR from the Autographa californica 
                     nucleopolyhedrovirus polyhedrin gene (Suzuki et al., 2006)"
     CDS             complement(3735..4323)
                     /codon_start=2
                     /product="baculovirus capsid-associated protein"
                     /label=ORF1629
                     /note="required for viral replication"
                     /translation="IPKSSTTNLIADVLADTINRRRVAMAKSSSEATSNDEGWDDDDNR
                     PNKANTPDVKYVQALFNVFTSSQLYTNDSDERNTKAHNILNDVEPLLQNKTQTNIDKAR
                     LLLQDLASFVALSENPLDSPAIGSEKQPLFETNRNLFYKSIEDLIFKFRYKDAENHLIF
                     ALTYHPKDYKFNELLKYVQQLSVNQQRTESSA"
     protein_bind    complement(4332..4365)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     rep_origin      complement(4533..5120)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4612..4631)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5133..5924,5925..5993))
                     /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     5756..5775
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 aactgttaca ttgcaaacgt ggtttcgtgt gccaagtgtg aaaaccgatg tttaatcaag
       61 gctctgacgc atttctacaa ccacgactcc aagtgtgtgg gtgaagtcat gcatctttta
      121 atcaaatccc aagatgtgta taaaccacca aactgccaaa aaatgaaaac tgtcgacaag
      181 ctctgtccgt ttgctggcaa ctgcaagggt ctcaatccta tttgtaatta ttgaataata
      241 aaacaattat aaatgtcaaa tttgtttttt attaacgata caaaccaaac gcaacaagaa
      301 catttgtagt attatctata attgaaaacg cgtagttata atcgctgagg taatatttaa
      361 aatcattttc aaatgattca cagttaattt gcgacaatat aattttattt tcacataaac
      421 tagacgcctt gtcgtcttct tcttcgtatt ccttctcttt ttcatttttc tcttcataaa
      481 aattaacata gttattatcg tatccatata tgtatctatc gtatagagta aattttttgt
      541 tgtcataaat atatatgtct tttttaatgg ggtgtatagt accgctgcgc atagtttttc
      601 tgtaatttac aacagtgcta ttttctggta gttcttcgga gtgtgttgct ttaattatta
      661 aatttatata atcaatgaat ttgggatcgt cggttttgta caatatgttg ccggcatagt
      721 acgcagcttc ttctagttca attacaccat tttttagcag caccggatta acataacttt
      781 ccaaaatgtt gtacgaaccg ttaaacaaaa acagttcacc tcccttttct atactattgt
      841 ctgcgagcag ttgtttgttg ttaaaaataa cagccattgt aatgagacgc acaaactaat
      901 atcacaaact ggaaatgtct atcaatatat agttgctgat ggccggccta ttaatagtaa
      961 tcaattacgg ggtcattagt tcatagccca tatatggagt tccgcgttac ataacttacg
     1021 gtaaatggcc cgcctggctg accgcccaac gacccccgcc cattgacgtc aataatgacg
     1081 tatgttccca tagtaacgcc aatagggact ttccattgac gtcaatgggt ggagtattta
     1141 cggtaaactg cccacttggc agtacatcaa gtgtatcata tgccaagtcc gccccctatt
     1201 gacgtcaatg acggtaaatg gcccgcctgg cattatgccc agtacatgac cttacgggac
     1261 tttcctactt ggcagtacat ctacgtatta gtcatcgcta ttaccatgct gatgcggttt
     1321 tggcagtaca ccaatgggcg tggatagcgg tttgactcac ggggatttcc aagtctccac
     1381 cccattgacg tcaatgggag tttgttttgg caccaaaatc aacgggactt tccaaaatgt
     1441 cgtaataacc ccgccccgtt gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat
     1501 ataagcagac gtcgtttagt gaaccgtcag atcactagat gctttattgc ggtagtttat
     1561 cacagttaaa ttgctaacgc cagtctcgaa cttaacgtgc agaagttggt cgtgaggcac
     1621 tgggcaggta agtatcgggc cctttgtgcg gggggagcgg ctcggggctg tccgcggggg
     1681 gacggctgcc ttcggggggg acggggcagg gcggggttcg gcttctggcg tgtgaccggc
     1741 ggctctagag cctctgctaa ccatgttcat gccttcttct ttttcctaca gctcctgggc
     1801 aacgtgctgg ttattgtgct gtctcatcat tttggcaaag aattggatcg gaccgaaatt
     1861 aatacgactc actatagggg aattgtgagc ggataacaat tccccggagt taatccggga
     1921 cctttaattc aacccaacac aatatattat agttaaataa gaattattat caaatcattt
     1981 gtatattaat taaaatacta tactgtaaat tacattttat ttacaatcaa aggagatata
     2041 ccatggtggg ccgcaacagc gcgattgcag ctggagtgtg cggcgcgctg ttcattggct
     2101 attgcattta ctttgatcgc aaacgccgca gcgatccgaa ctttaaatct agagtgagca
     2161 agggcgagga gctgttcacc ggggtggtgc ccatcctggt cgagctggac ggcgacgtaa
     2221 acggccacaa gttcagcgtg tccggcgagg gcgagggcga tgccacctac ggcaagctga
     2281 ccctgaagtt catcagcacc accggcaagc tgcccgtgcc ctggcccacc ctcgtgacca
     2341 ccctcggcta cggcctgcag tgcttcgccc gctaccccga ccacatgaag cagcacgact
     2401 tcttcaagtc cgccatgccc gaaggctacg tccaggagcg caccatcttc ttcaaggacg
     2461 acggcaacta caagacccgc gccgaggtga agttcgaggg cgacaccctg gtgaaccgca
     2521 tcgagctgaa gggcatcgac ttcaaggagg acggcaacat cctggggcac aagctggagt
     2581 acaactacaa cagccacaac gtctatatca ccgccgacaa gcagaagaac ggcatcaagg
     2641 caaacttcaa gatccgccac aacatcgagg acggcggcgt gcagctcgcc gaccactacc
     2701 agcagaacac ccccatcggc gacggccccg tgctgctgcc cgacaaccac tacctgagct
     2761 accagtccaa actgagcaaa gaccccaacg agaagcgcga tcacatggtc ctgctggagt
     2821 tcgtgaccgc cgccgggatc actctcggca tggacgagct gtacaagggt tctggaagtg
     2881 gatccatggt ggataacaaa ctggccacga ttgaccagtg gctgctgaaa aacgcgaaag
     2941 aagatgctat tgcagaactg aaaaaggctg gtatcaccca ggactatctg ttcaaccgta
     3001 tcaatgaggc gcattttgtg aatcaggtta acgtgcggaa gaacgtgatc ctgaaagctc
     3061 acgccggtgg ttctggtggt agccatcatc atcatcatca tggcggtgaa aatctgtatt
     3121 tccagggcgg ccgcgactac aaggacgacg acgacaaata aaagcttgcg gccgcacagc
     3181 tgtatacacg tgcaagccag ccagaactcg ccccggaaga ccccgaggat ctcgagcacc
     3241 accatcacca tcaccatcac taagtgatta acctcaggtg caggctgcct atcagaaggt
     3301 ggtggctggt gtggccaatg ccctggctca caaataccac tgagatcgat ctttttccct
     3361 ctgccaaaaa ttatggggac atcatgaagc cccttgagca tctgacttct ggctaataaa
     3421 ggaaatttat tttcattgca atagtgtgtt ggaatttttt gtgtctctca ctcggaagga
     3481 catatgggag ggcaaatcat ttaaaacatc agaatgagta tttggtttag agtttggcaa
     3541 catatgccca tatgtaacta gcataacccc ttggggcctc taaacgggtc ttgaggggtt
     3601 ttttgctgaa agcatgcgga ggaaattctc cttgaagttt ccctggtgtt caaagtaaag
     3661 gagtttgcac cagacgcacc tctgttcact ggtccggcgt attaaaacac gatacattgt
     3721 tattagtaca tttattaagc gctagattct gtgcgttgtt gatttacaga caattgttgt
     3781 acgtatttta ataattcatt aaatttataa tctttagggt ggtatgttag agcgaaaatc
     3841 aaatgatttt cagcgtcttt atatctgaat ttaaatatta aatcctcaat agatttgtaa
     3901 aataggtttc gattagtttc aaacaagggt tgtttttccg aaccgatggc tggactatct
     3961 aatggatttt cgctcaacgc cacaaaactt gccaaatctt gtagcagcaa tctagctttg
     4021 tcgatattcg tttgtgtttt gttttgtaat aaaggttcga cgtcgttcaa aatattatgc
     4081 gcttttgtat ttctttcatc actgtcgtta gtgtacaatt gactcgacgt aaacacgtta
     4141 aatagagctt ggacatattt aacatcgggc gtgttagctt tattaggccg attatcgtcg
     4201 tcgtcccaac cctcgtcgtt agaagttgct tccgaagacg attttgccat agccacacga
     4261 cgcctattaa ttgtgtcggc taacacgtcc gcgatcaaat ttgtagttga gctttttgga
     4321 attgcgatcg cataacttcg tatagcatac attatacgaa gttataagct cggaacgctg
     4381 cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta
     4441 tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc
     4501 aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc cctgacgagc
     4561 atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc
     4621 aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg
     4681 gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcaatgc tcacgctgta
     4741 ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg
     4801 ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac
     4861 acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag
     4921 gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat
     4981 ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat
     5041 ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc
     5101 gcagaaaaaa aggatctcaa gaagatcctt tgttaccaat gcttaatcag tgaggcacct
     5161 atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata
     5221 actacgatac gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca
     5281 cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga
     5341 agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga
     5401 gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg
     5461 gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga
     5521 gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt
     5581 gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct
     5641 cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca
     5701 ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat
     5761 accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga
     5821 aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc
     5881 aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg
     5941 caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc
     6001 ctttttcaat attattgaag catttatcag ggttattgtc tcatgtccgc gcgtt
//
