LOCUS       sequence-311740-       10066 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Targeted C-to-T in rice protoplasts.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 10066)
  TITLE     Gao Lab Plasmids
REFERENCE   2  (bases 1 to 10066)
  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"
COMMENT     Sequence Label: A3A-PBE-NG
FEATURES             Location/Qualifiers
     source          1..10066
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     RBS             6681..6689
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     CDS             6769..6816
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             6838..7086
                     /codon_start=1
                     /product="uracil-DNA glycosylase inhibitor from a Bacillus 
                     subtilis bacteriophage (Mol et al., 1995)"
                     /label=UGI
                     /translation="TNLSDIIEKETGKQLVIQESILMLPEEVEEVIGNKPESDILVHTA
                     YDESTDENVMLLTSDAPEYKPWALVIQDSNGENKIKML"
     CDS             7099..7119
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     polyA_signal    7177..7353
                     /label=CaMV poly(A) signal
                     /note="cauliflower mosaic virus polyadenylation signal"
     primer_bind     complement(7907..7923)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(7907..7923)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(7920..7942)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    7931..7947
                     /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(7955..7961,7962..7979,7980..7985))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    8000..8021
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(8138..8155)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(8309..8897)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(8389..8408)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(9068..9859,9860..9928))
                     /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     9691..9710
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(9929..10033)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 caagctaatt cgagctcggt acctgacccg gtcgtgcccc tctctagaga taatgagcat
       61 tgcatgtcta agttataaaa aattaccaca tatttttttt gtcacacttg tttgaagtgc
      121 agtttatcta tctttataca tatatttaaa ctttactcta cgaataatat aatctatagt
      181 actacaataa tatcagtgtt ttagagaatc atataaatga acagttagac atggtctaaa
      241 ggacaattga gtattttgac aacaggactc tacagtttta tctttttagt gtgcatgtgt
      301 tctccttttt ttttgcaaat agcttcacct atataatact tcatccattt tattagtaca
      361 tccatttagg gtttagggtt aatggttttt atagactaat ttttttagta catctatttt
      421 attctatttt agcctctaaa ttaagaaaac taaaactcta ttttagtttt tttatttaat
      481 aatttagata taaaatagaa taaaataaag tgactaaaaa ttaaacaaat accctttaag
      541 aaattaaaaa aactaaggaa acatttttct tgtttcgagt agataatgcc agcctgttaa
      601 acgccgtcga cgagtctaac ggacaccaac cagcgaacca gcagcgtcgc gtcgggccaa
      661 gcgaagcaga cggcacggca tctctgtcgc tgcctctgga cccctctcga tcgagagttc
      721 cgctccaccg ttggacttgc tccgctgtcg gcatccagaa attgcgtggc ggagcggcag
      781 acgtgagccg gcacggcagg cggcctcctc ctcctctcac ggcaccggca gctacggggg
      841 attcctttcc caccgctcct tcgctttccc ttcctcgccc gccgtaataa atagacaccc
      901 cctccacacc ctctttcccc aacctcgtgt tgttcggagc gcacacacac acaaccagat
      961 ctcccccaaa tccacccgtc ggcacctccg cttcaaggta cgccgctcgt cctccccccc
     1021 cccccctctc taccttctct agatcggcgt tccggtccat ggttagggcc cggtagttct
     1081 acttctgttc atgtttgtgt tagatccgtg tttgtgttag atccgtgctg ctagcgttcg
     1141 tacacggatg cgacctgtac gtcagacacg ttctgattgc taacttgcca gtgtttctct
     1201 ttggggaatc ctgggatggc tctagccgtt ccgcagacgg gatcgatttc atgatttttt
     1261 ttgtttcgtt gcatagggtt tggtttgccc ttttccttta tttcaatata tgccgtgcac
     1321 ttgtttgtcg ggtcatcttt tcatgctttt ttttgtcttg gttgtgatga tgtggtctgg
     1381 ttgggcggtc gttctagatc ggagtagaat taattctgtt tcaaactacc tggtggattt
     1441 attaattttg gatctgtatg tgtgtgccat acatattcat agttacgaat tgaagatgat
     1501 ggatggaaat atcgatctag gataggtata catgttgatg cgggttttac tgatgcatat
     1561 acagagatgc tttttgttcg cttggttgtg atgatgtggt gtggttgggc ggtcgttcat
     1621 tcgttctaga tcggagtaga atactgtttc aaactacctg gtgtatttat taattttgga
     1681 actgtatgtg tgtgtcatac atcttcatag ttacgagttt aagatggatg gaaatatcga
     1741 tctaggatag gtatacatgt tgatgtgggt tttactgatg catatacatg atggcatatg
     1801 cagcatctat tcatatgctc taaccttgag tacctatcta ttataataaa caagtatgtt
     1861 ttataattat tttgatctta atatacttgg atgatggcat atgcagcagc tatatgtgga
     1921 tttttttagc cctgccttca tacgctattt atttgcttgg tactgtttct tttgtcgatg
     1981 ctcaccctgt tgtttggtgt tacttctgca aagcttatgg aggccagccc ggctagcggc
     2041 ccaaggcatc tcatggaccc gcacatcttc accagcaact tcaacaacgg catcggcagg
     2101 cacaagacct acttgtgcta cgaggtggag aggctcgaca acggaacctc cgtgaagatg
     2161 gaccaacaca gggggttcct ccacaaccaa gccaagaacc tcctctgcgg cttctacggc
     2221 aggcacgccg agttgaggtt cctcgacttg gtgccatccc tccaactcga tccagcccaa
     2281 atctaccgcg tgacctggtt catctcctgg tccccatgct tctcctgggg ttgcgccggc
     2341 gaggttcggg ctttcctcca agaaaacacc cacgtccgcc tccgcatttt cgccgccagg
     2401 atctatgatt acgaccctct ctacaaggag gccctccaga tgctgcggga cgccggtgct
     2461 caggtgagta tcatgaccta cgacgagttc aagcactgct gggacacctt cgttgaccac
     2521 cagggctgcc cattccaacc atgggacggt ctggatgaac acagccaagc cttgtccggc
     2581 aggctccggg ccatcctcca aaaccagggg aactccggga gcgagacgcc aggcacctcc
     2641 gagtcggcca ccccagaatc tcttaaggac aagaagtact cgatcggcct cgccattggg
     2701 actaactctg ttggctgggc cgtgatcacc gacgagtaca aggtgccctc aaagaagttc
     2761 aaggtcctgg gcaacaccga tcggcattcc atcaagaaga atctcattgg cgctctcctg
     2821 ttcgacagcg gcgagacggc tgaggctacg cggctcaagc gcaccgcccg caggcggtac
     2881 acgcgcagga agaatcgcat ctgctacctg caggagattt tctccaacga gatggcgaag
     2941 gttgacgatt ctttcttcca caggctggag gagtcattcc tcgtggagga ggataagaag
     3001 cacgagcggc atccaatctt cggcaacatt gtcgacgagg ttgcctacca cgagaagtac
     3061 cctacgatct accatctgcg gaagaagctc gtggactcca cagataaggc ggacctccgc
     3121 ctgatctacc tcgctctggc ccacatgatt aagttcaggg gccatttcct gatcgagggg
     3181 gatctcaacc cggacaatag cgatgttgac aagctgttca tccagctcgt gcagacgtac
     3241 aaccagctct tcgaggagaa ccccattaat gcgtcaggcg tcgacgcgaa ggctatcctg
     3301 tccgctaggc tctcgaagtc tcggcgcctc gagaacctga tcgcccagct gccgggcgag
     3361 aagaagaacg gcctgttcgg gaatctcatt gcgctcagcc tggggctcac gcccaacttc
     3421 aagtcgaatt tcgatctcgc tgaggacgcc aagctgcagc tctccaagga cacatacgac
     3481 gatgacctgg ataacctcct ggcccagatc ggcgatcagt acgcggacct gttcctcgct
     3541 gccaagaatc tgtcggacgc catcctcctg tctgatattc tcagggtgaa caccgagatt
     3601 acgaaggctc cgctctcagc ctccatgatc aagcgctacg acgagcacca tcaggatctg
     3661 accctcctga aggcgctggt caggcagcag ctccccgaga agtacaagga gatcttcttc
     3721 gatcagtcga agaacggcta cgctgggtac attgacggcg gggcctctca ggaggagttc
     3781 tacaagttca tcaagccgat tctggagaag atggacggca cggaggagct gctggtgaag
     3841 ctcaatcgcg aggacctcct gaggaagcag cggacattcg ataacggcag catcccacac
     3901 cagattcatc tcggggagct gcacgctatc ctgaggaggc aggaggactt ctaccctttc
     3961 ctcaaggata accgcgagaa gatcgagaag attctgactt tcaggatccc gtactacgtc
     4021 ggcccactcg ctaggggcaa ctcccgcttc gcttggatga cccgcaagtc agaggagacg
     4081 atcacgccgt ggaacttcga ggaggtggtc gacaagggcg ctagcgctca gtcgttcatc
     4141 gagaggatga cgaatttcga caagaacctg ccaaatgaga aggtgctccc taagcactcg
     4201 ctcctgtacg agtacttcac agtctacaac gagctgacta aggtgaagta tgtgaccgag
     4261 ggcatgagga agccggcttt cctgtctggg gagcagaaga aggccatcgt ggacctcctg
     4321 ttcaagacca accggaaggt cacggttaag cagctcaagg aggactactt caagaagatt
     4381 gagtgcttcg attcggtcga gatctctggc gttgaggacc gcttcaacgc ctccctgggg
     4441 acctaccacg atctcctgaa gatcattaag gataaggact tcctggacaa cgaggagaat
     4501 gaggatatcc tcgaggacat tgtgctgaca ctcactctgt tcgaggaccg ggagatgatc
     4561 gaggagcgcc tgaagactta cgcccatctc ttcgatgaca aggtcatgaa gcagctcaag
     4621 aggaggaggt acaccggctg ggggaggctg agcaggaagc tcatcaacgg cattcgggac
     4681 aagcagtccg ggaagacgat cctcgacttc ctgaagagcg atggcttcgc gaaccgcaat
     4741 ttcatgcagc tgattcacga tgacagcctc acattcaagg aggatatcca gaaggctcag
     4801 gtgagcggcc agggggactc gctgcacgag catatcgcga acctcgctgg ctcgccagct
     4861 atcaagaagg ggattctgca gaccgtgaag gttgtggacg agctggtgaa ggtcatgggc
     4921 aggcacaagc ctgagaacat cgtcattgag atggcccggg agaatcagac cacgcagaag
     4981 ggccagaaga actcacgcga gaggatgaag aggatcgagg agggcattaa ggagctgggg
     5041 tcccagatcc tcaaggagca cccggtggag aacacgcagc tgcagaatga gaagctctac
     5101 ctgtactacc tccagaatgg ccgcgatatg tatgtggacc aggagctgga tattaacagg
     5161 ctcagcgatt acgacgtcga tcatatcgtt ccacagtcat tcctgaagga tgactccatt
     5221 gacaacaagg tcctcaccag gtcggacaag aaccggggca agtctgataa tgttccttca
     5281 gaggaggtcg ttaagaagat gaagaactac tggcgccagc tcctgaatgc caagctgatc
     5341 acgcagcgga agttcgataa cctcacaaag gctgagaggg gcgggctctc tgagctggac
     5401 aaggcgggct tcatcaagag gcagctggtc gagacacggc agatcactaa gcacgttgcg
     5461 cagattctcg actcacggat gaacactaag tacgatgaga atgacaagct gatccgcgag
     5521 gtgaaggtca tcaccctgaa gtcaaagctc gtctccgact tcaggaagga tttccagttc
     5581 tacaaggttc gggagatcaa caattaccac catgcccatg acgcgtacct gaacgcggtg
     5641 gtcggcacag ctctgatcaa gaagtaccca aagctcgaga gcgagttcgt gtacggggac
     5701 tacaaggttt acgatgtgag gaagatgatc gccaagtcgg agcaggagat tggcaaggct
     5761 accgccaagt acttcttcta ctctaacatt atgaatttct tcaagacaga gatcactctg
     5821 gccaatggcg agatccggaa gcgccccctc atcgagacga acggcgagac gggggagatc
     5881 gtgtgggaca agggcaggga tttcgcgacc gtcaggaagg ttctctccat gccacaagtg
     5941 aatatcgtca agaagacaga ggtccagact ggcgggttct ctaaggagtc aattcggcct
     6001 aagcggaaca gcgacaagct catcgcccgc aagaaggact gggatccgaa gaagtacggc
     6061 gggttcgtca gccccactgt ggcctactcg gtcctggttg tggcgaaggt tgagaagggc
     6121 aagtccaaga agctcaagag cgtgaaggag ctgctgggga tcacgattat ggagcgctcc
     6181 agcttcgaga agaacccgat cgatttcctg gaggcgaagg gctacaagga ggtgaagaag
     6241 gacctgatca ttaagctccc caagtactca ctcttcgagc tggagaacgg caggaagcgg
     6301 atgctggctt ccgctcgctt cctgcagaag gggaacgagc tggctctgcc gtccaagtat
     6361 gtgaacttcc tctacctggc ctcccactac gagaagctca agggcagccc cgaggacaac
     6421 gagcagaagc agctgttcgt cgagcagcac aagcattacc tcgacgagat cattgagcag
     6481 atttccgagt tctccaagcg cgtgatcctg gccgacgcga atctggataa ggtcctctcc
     6541 gcgtacaaca agcaccgcga caagccaatc agggagcagg ctgagaatat cattcatctc
     6601 ttcaccctga cgaacctcgg cgcccctcgt gctttcaagt acttcgacac aactatcgat
     6661 cgcaaggtat acagaagcac taaggaggtc ctggacgcga ccctcatcca ccagtcgatt
     6721 accggcctct acgagacgcg catcgacctg tctcagctcg ggggcgacaa gcggccagcg
     6781 gcgacgaaga aggcggggca ggcgaagaag aagaagaccc gcgactccgg cggcagcacg
     6841 aacctctccg acatcatcga gaaggagacg ggcaagcagc tcgtgatcca ggagagcatc
     6901 ctcatgctgc cggaggaggt ggaggaggtc atcggcaaca agcccgagtc cgacatcctc
     6961 gtgcacaccg cctacgacga gtccacggac gagaacgtca tgctcctgac gagcgacgct
     7021 ccagagtaca agccatgggc tctcgtgatc caggacagca acggcgagaa caagatcaag
     7081 atgctgtccg gcggctcccc aaagaagaag cggaaggtgt aggaattcgg tacgctgaaa
     7141 tcaccagtct ctctctacaa atctatctct ctctattttc tccataaata atgtgtgagt
     7201 agtttcccga taagggaaat tagggttctt atagggtttc gctcatgtgt tgagcatata
     7261 agaaaccctt agtatgtatt tgtatttgta aaatacttct atcaataaaa tttctaattc
     7321 ctaaaaccaa aatccagtac taaaatccag atctcctaaa gtccctatag atctttgtcg
     7381 tgaatataaa ccagacacga gacgactaaa cctggagccc agacgccgtt cgaagctaga
     7441 agtaccgctt aggcaggagg ccgttaggga aaagatgcta aggcagggtt ggttacgttg
     7501 actcccccgt aggtttggtt taaatatgat gaagtggacg gaaggaagga ggaagacaag
     7561 gaaggataag gttgcaggcc ctgtgcaagg taagaagatg gaaatttgat agaggtacgc
     7621 tactatactt atactatacg ctaagggaat gcttgtattt ataccctata ccccctaata
     7681 accccttatc aatttaagaa ataatccgca taagcccccg cttaaaaatt ggtatcagag
     7741 ccatgaatag gtctatgacc aaaactcaag aggataaaac ctcaccaaaa tacgaaagag
     7801 ttcttaactc taaagataaa agatctttca agatcaaaac tagttccctc acaccggtga
     7861 cggggatcgc atgcgatatc tcgagatcta gcttggcgta atcatggtca tagctgtttc
     7921 ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt
     7981 gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc
     8041 ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg
     8101 ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct
     8161 cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca
     8221 cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga
     8281 accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc
     8341 acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg
     8401 cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat
     8461 acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt
     8521 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc
     8581 agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg
     8641 acttatcgcc actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg
     8701 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg
     8761 gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg
     8821 gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca
     8881 gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga
     8941 acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga
     9001 tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt
     9061 ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt
     9121 catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat
     9181 ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag
     9241 caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct
     9301 ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt
     9361 tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg
     9421 cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca
     9481 aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt
     9541 tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat
     9601 gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac
     9661 cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa
     9721 aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt
     9781 tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt
     9841 tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa
     9901 gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt
     9961 atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa
    10021 taggggttcc gcgcacattt ccccgaaaag tgccacctgc cagtgc
//
