LOCUS       sequence-368780-        6411 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  original vector to make all constructs.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6411)
  AUTHORS   Cho SI, Lee S, Mok YG, Lim K, Lee J, Lee JM, Chung E, Kim JS
  TITLE     Targeted A-to-G base editing in human mitochondrial DNA with 
            programmable deaminases.
  JOURNAL   Cell. 2022 May 12;185(10):1764-1776.e12. doi: 
            10.1016/j.cell.2022.03.039. Epub 2022 Apr 25.
  PUBMED    35472302
REFERENCE   2  (bases 1 to 6411)
  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: 
            "10.1016/j.cell.2022.03.039"; journalName: "Cell"; date: 
            "2022-05-12- 12"; volume: "185"; issue: "10"; pages: "1764-1776.e12"
COMMENT     Sequence Label: TALED_Right-stuffer-8e
FEATURES             Location/Qualifiers
     source          1..6411
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             885..962
                     /codon_start=1
                     /product="mitochondrial presequence of human cytochrome c 
                     oxidase subunit VIII (Rizutto et al., 1989)"
                     /label=COX8 presequence
                     /translation="MASVLTPLLLRGLTGSARRLPVPRAK"
     CDS             975..1040
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             1466..2140
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /translation="MASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKM
                     RLKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHS
                     TGA"
     CDS             1466..2140
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /note="mammalian codon-optimized"
                     /translation="MASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKM
                     RLKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHS
                     TGA"
     primer_bind     complement(1596..1614)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1864..1883
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     2058..2081
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     CDS             join(2182..2184,2185..2898)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             join(2182..2184,2185..2895)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(2224..2245)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2485..2504)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2832..2853
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     promoter        complement(3585..3603)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(3586..3603)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(3623..3640)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3629..3853
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     polyA_signal    4084..4205
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4121..4140)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4175..4194
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4254..4270)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4254..4270)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4267..4289)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4278..4294
                     /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        complement(join(4302..4308,4309..4326,4327..4332))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4347..4368
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4485..4502)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4656..5244)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4736..4755)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5415..6206,6207..6275))
                     /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     6038..6057
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6276..6380)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggac tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagatggcc tctgtcttga
      901 caccactcct gctcaggggc ctgactggat cagctaggcg gctccccgtt cctcgcgcaa
      961 aaatacacag cctggactat aaggaccatg acggtgatta caaagatcac gacatagatt
     1021 ataaggatga tgatgataag gggatccgaa ttcaagatct acgcacgctc ggctacagcc
     1081 agcagcaaca ggagaagatc aaaccgaagg ttcgttcgac agtggcgcag caccacgagg
     1141 cactggtcgg ccatgggttt acacacgcgc acatcgttgc gctcagccaa cacccggcag
     1201 cgttagggac cgtcgctgtc aagtatcagg acatgatcgc agcgttgcca gaggcgacac
     1261 acgaagcgat cgttggcgtc ggcaaacagt ggtccggcgc acgcgctctg gaggccttgc
     1321 tcacggtggc gggagagttg agaggtccac cgttacagtt ggacacaggc caacttctca
     1381 agattgcaaa acgtggcggc gtgaccgcag tggaggcagt gcatgcatgg cgcaatgcac
     1441 tgacgggtgc ccccctgagg agacgatggc ctcctccgag gacgtcatca aggagttcat
     1501 gcgcttcaag gtgcgcatgg agggctccgt gaacggccac gagttcgaga tcgagggcga
     1561 gggcgagggc cgcccctacg agggcaccca gaccgccaag ctgaaggtga ccaagggcgg
     1621 ccccctgccc ttcgcctggg acatcctgtc ccctcagttc cagtacggct ccaaggccta
     1681 cgtgaagcac cccgccgaca tccccgacta cttgaagctg tccttccccg agggcttcaa
     1741 gtgggagcgc gtgatgaact tcgaggacgg cggcgtggtg accgtgaccc aggactcctc
     1801 cctgcaggac ggcgagttca tctacaaggt gaagctgcgc ggcaccaact tcccctccga
     1861 cggccccgta atgcagaaga agaccatggg ctgggaggcc tccaccgagc ggatgtaccc
     1921 cgaggacggc gccctgaagg gcgagatcaa gatgaggctg aagctgaagg acggcggcca
     1981 ctacgacgcc gaggtcaaga ccacctacat ggccaagaag cccgtgcagc tgcccggcgc
     2041 ctacaagacc gacatcaagc tggacatcac ctcccacaac gaggactaca ccatcgtgga
     2101 acagtacgag cgcgccgagg gccgccactc caccggcgcc gaattcatga catcaattat
     2161 tatacatcgg aggaggatcc agtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc
     2221 atcctggtcg agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggc
     2281 gagggcgatg ccacctacgg caagctgacc ctgaagttca tctgcaccac cggcaagctg
     2341 cccgtgccct ggcccaccct cgtgaccacc ctgacctacg gcgtgcagtg cttcagccgc
     2401 taccccgacc acatgaagca gcacgacttc ttcaagtccg ccatgcccga aggctacgtc
     2461 caggagcgca ccatcttctt caaggacgac ggcaactaca agacccgcgc cgaggtgaag
     2521 ttcgagggcg acaccctggt gaaccgcatc gagctgaagg gcatcgactt caaggaggac
     2581 ggcaacatcc tggggcacaa gctggagtac aactacaaca gccacaacgt ctatatcatg
     2641 gccgacaagc agaagaacgg catcaaggtg aacttcaaga tccgccacaa catcgaggac
     2701 ggcagcgtgc agctcgccga ccactaccag cagaacaccc ccatcggcga cggccccgtg
     2761 ctgctgcccg acaaccacta cctgagcacc cagtccgccc tgagcaaaga ccccaacgag
     2821 aagcgcgatc acatggtcct gctggagttc gtgaccgccg ccgggatcac tctcggcatg
     2881 gacgagctgt acaagtaacg tctcccggtc tgactccgga acaggtggtg gcgattgcaa
     2941 gcaacggcgg cggcaaacag gctctagaga gcattgttgc ccagctctcc agacctgatc
     3001 cggcgctagc cgcgttgcta gtctcaggaa gcgaaactcc tggtacctca gagtccgcta
     3061 ctcccgaatc ctctgaggtg gagttttccc acgagtactg gatgagacat gccctgaccc
     3121 tggccaagag ggcacgggat gagagggagg tgcctgtggg agccgtgctg gtgctgaaca
     3181 atagagtgat cggcgagggc tggaacagag ccatcggcct gcacgaccca acagcccatg
     3241 ccgaaattat ggccctgaga cagggcggcc tggtcatgca gaactacaga ctgattgacg
     3301 ccaccctgta cgtgacattc gagccttgcg tgatgtgcgc cggcgccatg atccactcta
     3361 ggatcggccg cgtggtgttt ggcgtgagga actcaaaaag aggcgccgca ggctccctga
     3421 tgaacgtgct gaactacccc ggcatgaatc accgcgtcga aattaccgag ggaatcctgg
     3481 cagatgaatg tgccgccctg ctgtgcgatt tctatcggat gcctagacag gtgttcaatg
     3541 ctcagaagaa ggcccagagc tccatcaact agtcagggcc ctattctata gtgtcaccta
     3601 aatgctagag ctcgctgatc agcctcgact gtgccttcta gttgccagcc atctgttgtt
     3661 tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt cctttcctaa
     3721 taaaatgagg aaattgcatc gcattgtctg agtaggtgtc attctattct ggggggtggg
     3781 gtggggcagg acagcaaggg ggaggattgg gaagacaata gcaggcatgc tggggatgcg
     3841 gtgggctcta tggcttctga ggcggaaaga accagctggg gctctagggg gtatccccac
     3901 gctgcagtga gcgggactct ggggttcgaa atgaccgacc aagcgacgcc caacctgcca
     3961 tcacgagatt tcgattccac cgccgccttc tatgaaaggt tgggcttcgg aatcgttttc
     4021 cgggacgccg gctggatgat cctccagcgc ggggatctca tgctggagtt cttcgcccac
     4081 cccaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc
     4141 acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta
     4201 tcttatcatg tctgtatacc gtcgacctct agctagagct tggcgtaatc atggtcatag
     4261 ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc
     4321 ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc
     4381 tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa
     4441 cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg
     4501 ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg
     4561 ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag
     4621 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
     4681 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
     4741 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
     4801 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
     4861 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     4921 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
     4981 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
     5041 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca
     5101 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
     5161 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
     5221 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
     5281 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc
     5341 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa
     5401 acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
     5461 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
     5521 ttaccatctg gccccagtgc tgcaatgata ccgcgagatc cacgctcacc ggctccagat
     5581 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
     5641 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
     5701 aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt
     5761 ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg
     5821 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc
     5881 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc
     5941 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg
     6001 cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga
     6061 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta
     6121 ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct
     6181 tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag
     6241 ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca atattattga
     6301 agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat
     6361 aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgt c
//
