LOCUS       sequence-358084-        9563 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Dual expression construct encoding shRNA-resistant mtIF3 and mCherry
            from separate promoters.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9563)
  AUTHORS   Lee S, Park D, Lim C, Kim JI, Min KT
  TITLE     mtIF3 is locally translated in axons and regulates mitochondrial 
            translation for axonal growth.
  JOURNAL   BMC Biol. 2022 Jan 7;20(1):12. doi: 10.1186/s12915-021-01215-w.
  PUBMED    34996455
REFERENCE   2  (bases 1 to 9563)
  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: "BMC Biol.";
            date: "2022-01-7"; volume: "20(1):12. doi"; pages: " 
            10.1186/s12915-021-01215-w"
COMMENT     Sequence Label: 
            psiCheck2-SV40p-5'UTR-mtIF3(SR)-3xFLAG-3'UTR-HSVTKp-mCherry
FEATURES             Location/Qualifiers
     source          1..9563
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        62..419
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        62..253
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(85..105)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      270..405
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     intron          489..621
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     604..623
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        666..684
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1766..1831
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             5869..5886
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     polyA_signal    5918..5966
                     /note="synthetic polyadenylation signal"
     promoter        5975..6726
                     /label=HSV TK promoter
                     /note="herpes simplex virus thymidine kinase promoter"
     CDS             6768..7478
                     /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(6913..6931)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     7181..7200
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    7518..7639
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7529..7548)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     7583..7602
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        7772..7876
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(7877..7945,7946..8737)
                     /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(8095..8114)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      8908..9496
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     9397..9416
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 agatctgcgc agcaccatgg cctgaaataa cctctgaaag aggaacttgg ttaggtacct
       61 tctgaggcgg aaagaaccag ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag
      121 gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg
      181 gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag
      241 caaccatagt cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc
      301 attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctcgg
      361 cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa
      421 agcttgattc ttctgacaca acagtctcga acttaagctg cagaagttgg tcgtgaggca
      481 ctgggcaggt aagtatcaag gttacaagac aggtttaagg agaccaatag aaactgggct
      541 tgtcgagaca gagaagactc ttgcgtttct gataggcacc tattggtctt actgacatcc
      601 actttgcctt tctctccaca ggtgtccact cccagttcaa ttacagctct taaggctaga
      661 gtacttaata cgactcacta taggctagcg gaagccgacg cggtgctccg gcggagtcca
      721 ggtacggacc tgctagttct tcctgctcgc ctaccccggc ccagggtctc actatttatc
      781 tctggctgtc ctgaaacgca ctcggtagac cagtctgacc tcaaacctat agaaatcctc
      841 ctgcctctac ctccgaagtg ctgggattaa aggcgtgcac cgctcgccgt gtcaggactt
      901 ttgaaattcc ttccggaatt gaaggggtac catggctgtt cttctgatga ggttgatgct
      961 acaaactaca aagcttgacc ataacttaat tggaagatgt ttgcaaagac acgctgtgaa
     1021 gccagatcca gcacagctgt ccctgtctgc gtccacccca aagctgttgt acctgacttc
     1081 tgcaaaaggt tttagtactg ctggagaccc acagggtgag aggaaacaga aaagaaggga
     1141 tgctttctct aacaccggcc ggaagattag tgagcggatt atccgcgtcc tggatgagaa
     1201 gggcatggac ctggggatga tgcatcgagc agatgtgatc agactcatga acaagcagga
     1261 cctgaggctg gtgcagagga acaccagctc agagccaccg gagtaccagc ttatgaccgg
     1321 agagcagatc caccaggagc ggctaaagct ccgggaacag gaaaaggcca aacccaaaac
     1381 tggaccaact atgacgaaag agctcgtttt ctcttcaaat attggacagc acgatttaga
     1441 caccaagagt aaacagatcc agcagtggat agagaagaaa tatcatgtac aggtgaccat
     1501 taagaggagg aaagatgcgg agcagtcaga ggaggaaacg gaggagatat ttaaccagat
     1561 tctccagact atgcctgaca tagcaacctt ctcaagcaga ccaaaagcca tcagaggagg
     1621 gactgcttcc atgtgtgttt ttcgtcactt gagcaagaaa gaagaaaagg cgtacagaga
     1681 atcccaggag agccagagaa gagacacttt gagcaaagac gacgacggaa acagcaaaga
     1741 gtcggatgtt gtgtgtcaga ccggtgacta taaggaccac gacggagact acaaggatca
     1801 tgatattgat tacaaagacg atgacgataa gtaagcggcc gcctttaata ggaaaagtgc
     1861 aggccaggag cctctgctgg ccgccctgaa gtcttcccgc ctagcagaca gtgagtaagg
     1921 cggctggcgg gggcagctgg gcctctgggc tgctgctgga ctctgcgctg gcgttaggca
     1981 aggagtagtc gggctcttgt ttcctcttag gagggaagta cccactgagg cgagaggcca
     2041 agctgcgtct ctcccgaggg gctcttacct tcaggagtga ggaagacaga caactgagtt
     2101 gcccattaca caacggggat ccactccaac agatgagccc ctgatgggag tcacaccctc
     2161 agcaggcaca gacacgcagg atggggtatg cggatggaca gacgccaagc aggcctcaca
     2221 gccgagtgct ggccccagca gctgccactt actttgagct tcatcctctt cttgtcggga
     2281 tcgtgcacga cactgtgcct catgagactc tgctgcagtg caagggaaag aaggacgctt
     2341 gacctgggcc tgcagacgtg gctcagtgct tacagcattc gctgctcgag gaggacacgg
     2401 gtttggttcc cagcacccgt actccagttc cagtaaaaac tcttggtgtc tgctaacctt
     2461 aagatgcagt ggtccagtgt gcaataataa actaccagct gcctttctgt tgtgtagtga
     2521 atctgacctt cacctgactc cctaaccaga gccaactcct tggcagagcg agcgactgtc
     2581 agcctgctga cacgcatggg atttacttct tcccggtcct cctgagaagg gaaagccggc
     2641 tatgcttgaa catccaggct catgctgggg taactgaatc tacctcagct gcgtccctaa
     2701 gtgccagggg gaaatctgac ctctgccctg ctggctatgc tgacagtgct gaccacggga
     2761 cctttgctgt gggctggtct gggttatgtt cagtgacttt gtaatccaca gtaagagctc
     2821 tttagaaacc agctaggctt aggcaaactg ttcagttccc tactaggaac catgttccca
     2881 cgcgatccat gaatcatgcc aagattccag agtctagagg ctgtgcattt gtgctaccca
     2941 cctcctctcc cagaccctag gcgacggtca gaagctccta atcacctcag aacatcctct
     3001 aagtaggaat acaggccccg ggctctccca ctcagcatgg ctagaatccc ccaggagtgg
     3061 ttctcaacct gctactcatg actcctctgg ggtcatatac cagatcgcct gaatatcatt
     3121 atttacaatc cataccagta gcaaaattat agttatgaag tagcaactaa ataatattat
     3181 ggttgggggt ccccacaaca tgaggaactg ttattgggtc ccagtattag gaaggttgag
     3241 aaccactgct ctagaggtcc taaataggcc tgtttagcct cccaacccca gggctcctgc
     3301 atttccaagt tctgcaggct ggacatgaac acaccagcct ggagcgggag acactcactt
     3361 tcattgcgaa tgtcttgcca cagccagcgt gctcacacac aaatgggcgc ttttcctcgt
     3421 ggaaggagag aatgtggctc tgcaggttga acacggttgt gtaggttctt ccgcagcctt
     3481 gccgcggaca gcggtacaca tccctttccg gggcgtgagt cttcatgtgc tgcttaaggt
     3541 aatctctgcg cttgaacatc ctctgacata cattgcaggt tatgtcctct gggaggagaa
     3601 acacacacgt gtaaccagcg tcccatgagt aacccgtctg gtcctatgac tacacacaat
     3661 ctggggactt cacatgggaa atgtcagctt gtctcactgc tatgctggtt tcaaattcct
     3721 caagttcact catggctccc aacatcctga gtcctcgcct gtcaaggagc aggcgtcctc
     3781 ggtgtgataa ctgagcacga ggaagtatgc cctcatagca tctaccttta ccctctgaca
     3841 cagtagcatc tgctaagtcc tgcagggcct cacgaacacc cgggcacagc cgcaccagct
     3901 ctgaggcctg cagggctccc tcgactgtca cagcccagga agccagcact accctgaact
     3961 cgatgcagag cccccttcct ttacagcgca agtgggtatc tcactctctc agagtacacc
     4021 atgcactccc cagagctcag agcacacaca ccatgggctc cccggggttc agagcacacc
     4081 gggctctccc cagggccgat gcgcttccac agctcctgcc taccttacct ttatgggctt
     4141 ctctcatgtg tttcaggagc tctgtccacg tttttcccat gaaagaacat cccttttgac
     4201 acaggtagcc tgttcaaaac aaaaacaaaa caagttaccc acagcctgtt gaatgcccag
     4261 ggcaatccct aatgacagcc aagaaagccc gtgccaagtg cactcccctc tctgaagtgt
     4321 gctgctctgc tgggtgcaat gcagcgggct ccagtggccc ctgctctgaa agctcccatc
     4381 agccagactt ttgcacactt gagtggcaca taaccaggta gcgtaggaac agtatagcac
     4441 cagtggacgt ggcaaagagg atagcggcaa aacacacagc acagtgtctc ttggcataag
     4501 aggcaaacac tcaaataaca caacaggcgc ttgcttgtca cagaactttg tctactcagc
     4561 gtgactacag aatagaccca gcctaccctg ccggtggcat gttcctcgcg gccggcacag
     4621 tccccatcta cgtaccctcg tgaactttcc catgccgttt cagcctgctg ggcgaggcaa
     4681 agtgcttccc gcatccctcg tgggtacacc taggacaata gcaggcaaag catcagccag
     4741 gccctttcca tggacaattc taagattccc tcttctccca caccaaagcc catgtcttcc
     4801 cacaccacac ctgtcatcca aaacaaagag taagtacagt cccgaatatg gacatagtgg
     4861 cttggccagc ttgggagcaa aacaagccat cctgagaaaa agtcacatct tttgtttgtg
     4921 ttatcaaact aaaagttctc tgaattctgg aaatgtggct acagaagcga catcaagagg
     4981 ctgtctgact cggtttgcac tggaccagac acagagctgt tgtctggggt ctgcccaata
     5041 tgctgcatac ggtcagatga gaacacagca cagaggctgg ggaccagcac cagaaacaca
     5101 cgagaatgta cagcagtcgg tgcagagctc caggcactgg cagcacaaac aggccacagg
     5161 gctgctggta agacctcggt cactgtagct gcaccagtgc aaagagcagc ctatgggagc
     5221 caggggcaag gccccatcac ccggactgtg cagggagaca ctgccctagg agggaggcag
     5281 gcaggtaacc tggcacccct gcccacctcc tctttgtccc tcttgagtac tcaggaatcc
     5341 ctcaaaacaa gcagggcctc attactcctg ttctatagta agaaaaaaaa aagccctcat
     5401 gacacagagc cttcctcggg atcctgatct tagcgttcta ctctgaatcc cagagcgctg
     5461 gcaaagcacc aacctgaaga gcggctcgct ggtgtgctgg cactgatggg ttctcagctg
     5521 ctggtgcttc ttaaaggcct tcttgcaacc ctcataactg cactgtttaa aaaaaataaa
     5581 tgcacggttt agtacaggca ggggatcatc agttgctgaa gtcctttaaa aggcaggaga
     5641 gattgctcaa tggttaggga cacttgcttc tctggtccca ggtttggact gcagcaccca
     5701 cgtggtgcct cacaaccatc tgactccagt tccacaggac ctcacaccct cttctgacct
     5761 ccacaggcac cgagcacgca tatggtatat agatacatat gcaggcaaaa cgcttgcaca
     5821 taaaaccttt tttaactaaa caataaagtc tctctggatc tgaccggtca tcatcaccat
     5881 caccattgag tttaaaccta gagcggccgc tggccgcaat aaaatatctt tattttcatt
     5941 acatctgtgt gttggttttt tgtgtgagga tctaaatgag tcttcggacc tcgcgggggc
     6001 cgcttaagcg gtggttaggg tttgtctgac gcggggggag ggggaaggaa cgaaacactc
     6061 tcattcggag gcggctcggg gtttggtctt ggtggccacg ggcacgcaga agagcgccgc
     6121 gatcctctta agcacccccc cgccctccgt ggaggcgggg gtttggtcgg cgggtggtaa
     6181 ctggcgggcc gctgactcgg gcgggtcgcg cgccccagag tgtgaccttt tcggtctgct
     6241 cgcagacccc cgggcggcgc cgccgcggcg gcgacgggct cgctgggtcc taggctccat
     6301 ggggaccgta tacgtggaca ggctctggag catccgcacg actgcggtga tattaccgga
     6361 gaccttctgc gggacgagcc gggtcacgcg gctgacgcgg agcgtccgtt gggcgacaaa
     6421 caccaggacg gggcacaggt acactatctt gtcacccgga ggcgcgaggg actgcaggag
     6481 cttcagggag tggcgcagct gcttcatccc cgtggcccgt tgctcgcgtt tgctggcggt
     6541 gtccccggaa gaaatatatt tgcatgtctt tagttctatg atgacacaaa ccccgcccag
     6601 cgtcttgtca ttggcgaatt cgaacacgca gatgcagtcg gggcggcgcg gtcccaggtc
     6661 cacttcgcat attaaggtga cgcgtgtggc ctcgaacacc gagcgaccct gcagcgaccc
     6721 gcttaaaagc ttggcattcc ggtactgttg gtaaagccac cgggcccatg gtgagcaagg
     6781 gcgaggagga taacatggcc atcatcaagg agttcatgcg cttcaaggtg cacatggagg
     6841 gctccgtgaa cggccacgag ttcgagatcg agggcgaggg cgagggccgc ccctacgagg
     6901 gcacccagac cgccaagctg aaggtgacca agggtggccc cctgcccttc gcctgggaca
     6961 tcctgtcccc tcagttcatg tacggctcca aggcctacgt gaagcacccc gccgacatcc
     7021 ccgactactt gaagctgtcc ttccccgagg gcttcaagtg ggagcgcgtg atgaacttcg
     7081 aggacggcgg cgtggtgacc gtgacccagg actcctccct gcaggacggc gagttcatct
     7141 acaaggtgaa gctgcgcggc accaacttcc cctccgacgg ccccgtaatg cagaagaaga
     7201 ccatgggctg ggaggcctcc tccgagcgga tgtaccccga ggacggcgcc ctgaagggcg
     7261 agatcaagca gaggctgaag ctgaaggacg gcggccacta cgacgctgag gtcaagacca
     7321 cctacaaggc caagaagccc gtgcagctgc ccggcgccta caacgtcaac atcaagttgg
     7381 acatcacctc ccacaacgag gactacacca tcgtggaaca gtacgaacgc gccgagggcc
     7441 gccactccac cggcggcatg gacgagctgt acaagtagtc tagagtcggg gcggccggcc
     7501 gcttcgagca gacatgataa gatacattga tgagtttgga caaaccacaa ctagaatgca
     7561 gtgaaaaaaa tgctttattt gtgaaatttg tgatgctatt gctttatttg taaccattat
     7621 aagctgcaat aaacaagtta acaacaacaa ttgcattcat tttatgtttc aggttcaggg
     7681 ggaggtgtgg gaggtttttt aaagcaagta aaacctctac aaatgtggta aaatcgataa
     7741 ggatccaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta
     7801 aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata
     7861 ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc
     7921 ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga
     7981 agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct
     8041 tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg
     8101 tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta
     8161 ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat
     8221 gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt
     8281 acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga
     8341 tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga
     8401 gcgtgacacc acgatgcctg tagcaatggc aacaacgttg cgcaaactat taactggcga
     8461 actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc
     8521 aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc
     8581 cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg
     8641 tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat
     8701 cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata
     8761 tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct
     8821 ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga
     8881 ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg
     8941 cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc
     9001 aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct
     9061 agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc
     9121 tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt
     9181 ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg
     9241 cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct
     9301 atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag
     9361 ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag
     9421 tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg
     9481 gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg
     9541 gccttttgct cacatggctc gac
//
