LOCUS       sequence-368781-        6270 bp    DNA     circular SYN 14-APR-2025
DEFINITION  original vector to make all constructs.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6270)
  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 6270)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 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-1397N
FEATURES             Location/Qualifiers
     source          1..6270
                     /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(3444..3462)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(3445..3462)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(3482..3499)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3488..3712
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     polyA_signal    3943..4064
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3980..3999)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4034..4053
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4113..4129)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4113..4129)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4126..4148)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4137..4153
                     /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(4161..4167,4168..4185,4186..4191))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4206..4227
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4344..4361)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4515..5103)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4595..4614)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5274..6065,6066..6134))
                     /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     5897..5916
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6135..6239)
                     /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 cgcgttgaca aatgatcatc ttgttgctct tgcttgtctt ggcggacgac
     3061 ctgccctgga tgctgtgaaa aaggggttgg ggggctctgg atcaggtagc tacgcacttg
     3121 gtccttacca gattagcgca ccccaactcc ccgcctataa tggtcaaacc gtcgggacct
     3181 tttactacgt aaacgatgct ggtgggctgg aatccaaagt attctcctca gggggcccta
     3241 caccctaccc caactacgcc aatgctggtc atgtagaagg gcagtcagca ctgtttatgc
     3301 gcgataatgg tataagcgag gggttggtct tccataacaa cccagagggt acttgtggct
     3361 tctgtgtgaa tatgactgaa acccttctgc ccgaaaatgc caagatgact gtcgtcccac
     3421 ctgaaggcta gtcagggccc tattctatag tgtcacctaa atgctagagc tcgctgatca
     3481 gcctcgactg tgccttctag ttgccagcca tctgttgttt gcccctcccc cgtgccttcc
     3541 ttgaccctgg aaggtgccac tcccactgtc ctttcctaat aaaatgagga aattgcatcg
     3601 cattgtctga gtaggtgtca ttctattctg gggggtgggg tggggcagga cagcaagggg
     3661 gaggattggg aagacaatag caggcatgct ggggatgcgg tgggctctat ggcttctgag
     3721 gcggaaagaa ccagctgggg ctctaggggg tatccccacg ctgcagtgag cgggactctg
     3781 gggttcgaaa tgaccgacca agcgacgccc aacctgccat cacgagattt cgattccacc
     3841 gccgccttct atgaaaggtt gggcttcgga atcgttttcc gggacgccgg ctggatgatc
     3901 ctccagcgcg gggatctcat gctggagttc ttcgcccacc ccaacttgtt tattgcagct
     3961 tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc atttttttca
     4021 ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctgtataccg
     4081 tcgacctcta gctagagctt ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt
     4141 tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa agcctggggt
     4201 gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg
     4261 ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg
     4321 cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg
     4381 cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat
     4441 aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc
     4501 gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc
     4561 tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga
     4621 agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt
     4681 ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg
     4741 taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc
     4801 gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg
     4861 gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc
     4921 ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg
     4981 ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc
     5041 gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct
     5101 caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt
     5161 taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa
     5221 aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa
     5281 tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc
     5341 tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct
     5401 gcaatgatac cgcgagatcc acgctcaccg gctccagatt tatcagcaat aaaccagcca
     5461 gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt
     5521 aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt
     5581 gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc
     5641 ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc
     5701 tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt
     5761 atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact
     5821 ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc
     5881 ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt
     5941 ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg
     6001 atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct
     6061 gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa
     6121 tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt
     6181 ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc
     6241 acatttcccc gaaaagtgcc acctgacgtc
//
