LOCUS       sequence-404880-        6211 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For DdCBE editing using Ddd9 at JAK2 site in human cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6211)
  AUTHORS   Huang J, Lin Q, Fei H, He Z, Xu H, Li Y, Qu K, Han P, Gao Q, Li B, 
            Liu G, Zhang L, Hu J, Zhang R, Zuo E, Luo Y, Ran Y, Qiu JL, Zhao KT,
            Gao C
  TITLE     Discovery of deaminase functions by structure-based protein 
            clustering.
  JOURNAL   Cell. 2023 Jul 20;186(15):3182-3195.e14. doi: 
            10.1016/j.cell.2023.05.041. Epub 2023 Jun 27.
  PUBMED    37379837
REFERENCE   2  (bases 1 to 6211)
  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.2023.05.041"; journalName: "Cell"; date: 
            "2023-07-20- 20"; volume: "186"; issue: "15"; pages: "3182-3195.e14"
COMMENT     Sequence Label: pCMV-TALE-R-JAK2-Ddd9-C
FEATURES             Location/Qualifiers
     source          1..6211
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        132..335
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     286..306
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     377..396
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        377..395
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             421..486
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             493..513
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2929..3177
                     /codon_start=1
                     /product="uracil-DNA glycosylase inhibitor from a Bacillus 
                     subtilis bacteriophage (Mol et al., 1995)"
                     /label=UGI
                     /translation="TNLSDIIEKETGKQLVIQESILMLPEEVEEVIGNKPESDILVHTA
                     YDESTDENVMLLTSDAPEYKPWALVIQDSNGENKIKML"
     CDS             3220..3240
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(3266..3283)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3272..3496
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(3567..3583)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3567..3583)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3580..3602)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3591..3607
                     /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(3615..3621,3622..3639,3640..3645))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3660..3681
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3798..3815)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3969..4557)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4049..4068)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4728..5519,5520..5588))
                     /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     5351..5370
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5589..5693)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(5772..5791)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        join(5963..6211,1..131)
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 atatgccaag tacgccccct attgacgtca atgacggtaa atggcccgcc tggcattatg
       61 cccagtacat gaccttatgg gactttccta cttggcagta catctacgta ttagtcatcg
      121 ctattaccat ggtgatgcgg ttttggcagt acatcaatgg gcgtggatag cggtttgact
      181 cacggggatt tccaagtctc caccccattg acgtcaatgg gagtttgttt tggcaccaaa
      241 atcaacggga ctttccaaaa tgtcgtaaca actccgcccc attgacgcaa atgggcggta
      301 ggcgtgtacg gtgggaggtc tatataagca gagctggttt agtgaaccgt cagatccgct
      361 agagatccgc ggccgctaat acgactcact atagggagac ccaagctggc tagcaccatg
      421 gattacaagg accacgacgg cgattacaag gaccacgaca tcgactacaa agacgacgac
      481 gacaagatgg cccctaagaa aaagaggaag gtgggcatcc acagaggggt acctatgatg
      541 gtggacctgc ggaccctggg ttatagccag cagcagcagg agaagatcaa gcccaaggtg
      601 cggagcaccg ttgcccagca ccacgaggcg ctggtcggac acggattcac ccacgcccac
      661 atcgtggccc tatctcagca tcctgctgcc ctcggcaccg tggccgtgaa gtaccaggat
      721 atgatcgctg ctctcccaga ggccacacac gaggccatcg tgggagtggg caagcagtgg
      781 tccggcgcca gagccttgga agccctgctg accgtggctg gcgagctgag aggacctcct
      841 ctgcagctgg acaccggcca gctgctgaag atcgccaagc ggggcggcgt gaccgccgtg
      901 gaagccgtgc acgcttggcg gaacgccctg acaggcgccc ctctgaacct gacccccgac
      961 caggtggtgg ccatcgccag ccacgacggc gggaagcagg ccctagagac tgtgcagaga
     1021 ttgctgcctg tgctgtgcca ggaccacggc ctgaccccag aacaagtcgt ggctatcgcc
     1081 agcaacggcg gcggcaagca ggcgttggaa acagtgcaga gactgctgcc cgtgctgtgt
     1141 caggcccacg gcctgacccc cgatcaggtc gtggccattg ccaacaacaa cggcggaaag
     1201 caggccctag aaaccgtgca gagactgctg cccgtgctgt gtcaggccca cggcctgaca
     1261 cctgcccagg tggttgctat cgcctccaac atcggaggaa agcaggctct ggaaaccgtg
     1321 cagagactgc tgcctgtgct gtgccaggat catggactga caccggatca ggtggtggcc
     1381 atagcatcta acatcggcgg caagcaagcc ctggagacag tacaaagact tctgcctgtg
     1441 ctgtgccagg accacggcct cacacccgag caggtggtgg ccattgcgtc caatatcggc
     1501 ggaaagcagg ccctggagac cgtgcaacga ctgctcccag tcctgtgcca ggctcacgga
     1561 ctcacccccg atcaggtggt ggctatagcc tctaacatcg gcggcaagca ggccctggag
     1621 acagtgcaaa gactgctgcc cgtgctgtgc caggctcacg gtctgacccc tgctcaggtg
     1681 gtcgccatcg cctctaacat cggaggcaag caggctttag agaccgttca aagactcctg
     1741 cccgtgctgt gtcaggatca cggcctgacc cctgatcagg tcgtggccat cgcaaacaac
     1801 aacggaggca agcaggcact ggaaaccgtg cagagactgc ttcccgtact gtgccaggac
     1861 cacggcctga cccctgaaca ggtggttgcc atcgccagca acatcggcgg aaagcaggcc
     1921 ctggagacgg tgcaaaggct gctgcctgtg ctgtgccagg cccacggcct gacccctgac
     1981 caggtagtgg ccatcgccag ccacgacgga gggaagcagg ctctggagac agtgcagaga
     2041 ctgctgcctg tgctgtgcca ggcccacggc ctgacccctg ctcaggtggt ggccatcgcc
     2101 tccaatggtg gcggcaaaca agctctggag acagtgcagc ggcttctgcc tgtgctgtgc
     2161 caggaccacg gcctgacccc tgaccaggtg gtggctatcg cctcccacga tggcggcaaa
     2221 caggccttag agaccgtgca acggctgctg cctgtgctgt gccaggatca cgggctgacc
     2281 cccgagcaag tggtggccat cgcaagcaac ggcggcggca aacaggcact tgaaacagtg
     2341 cagcgcctgc tgcctgtgct gtgccaagcc cacggcctga cccctgatca agtggttgct
     2401 atcgccaata ataacggcgg caaacaggcc ctggaaacag tgcagcgcct gctgccagtg
     2461 ctgtgtcagg cccacggcct gacaccagca caagtggtgg ccatcgccag ccacgacggc
     2521 ggcaaacagg ccctggaaac agtgcagaga ctgctgcctg ttctgtgtca ggaccacgga
     2581 ctgacccctg agcaggtggt ggccatcgcc tctaacatcg gcggaaggcc tgccctggag
     2641 agcatcgtgg cccagctgag caggcctgat cctgctctgg ccgccctgac caacgaccac
     2701 ctggtcgccc tggcctgtct gggcggccgg cccgccctgg acgccgtgaa gaagggcctg
     2761 ccccacgccc ctgccctgat caagcggaca aacagaagaa ttcctgaaag aaccagccat
     2821 agagtggcag gatccacagt gacccccagc agccgctggt cggcctctag aaccttcaca
     2881 ggaaatgatc gggaccccaa gccgtggcca agaggctccg gaggaagcac caacctgtct
     2941 gacatcatcg agaaggagac aggcaagcag ctggtcatcc aggagagcat cctgatgctg
     3001 cccgaagaag tcgaagaagt gatcggaaac aagcctgaga gcgatatcct ggtccatacc
     3061 gcctacgacg agagtaccga cgaaaatgtg atgctgctga catccgacgc cccagagtat
     3121 aagccctggg ctctggtcat ccaggattcc aacggagaga acaaaatcaa aatgctgtct
     3181 ggcggctcaa aaagaaccgc cgacggcagc gaattcgagc ccaagaagaa gaggaaagtc
     3241 taatgagttt aaacccgctg atcagcctcg actgtgcctt ctagttgcca gccatctgtt
     3301 gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg ccactcccac tgtcctttcc
     3361 taataaaatg aggaaattgc atcgcattgt ctgagtaggt gtcattctat tctggggggt
     3421 ggggtggggc aggacagcaa gggggaggat tgggaagaca atagcaggca tgctggggat
     3481 gcggtgggct ctatggcttc tgaggcggaa agaaccagct ggggctcgat accgtcgacc
     3541 tctagctaga gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg
     3601 ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagccta gggtgcctaa
     3661 tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac
     3721 ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt
     3781 gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga
     3841 gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca
     3901 ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg
     3961 ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt
     4021 cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc
     4081 ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct
     4141 tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc
     4201 gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta
     4261 tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca
     4321 gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag
     4381 tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag
     4441 ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt
     4501 agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa
     4561 gatcctttga tcttttctac ggggtctgac actcagtgga acgaaaactc acgttaaggg
     4621 attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga
     4681 agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta
     4741 atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc
     4801 cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg
     4861 ataccgcgag acccacgctc accggctcca gatttatcag caataaacca gccagccgga
     4921 agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt
     4981 tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt
     5041 gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc
     5101 caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc
     5161 ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca
     5221 gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag
     5281 tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg
     5341 tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa
     5401 cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa
     5461 cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga
     5521 gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga
     5581 atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg
     5641 agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt
     5701 ccccgaaaag tgccacctga cgtcgacgga tcgggagatc gatctcccga tcccctaggg
     5761 tcgactctca gtacaatctg ctctgatgcc gcatagttaa gccagtatct gctccctgct
     5821 tgtgtgttgg aggtcgctga gtagtgcgcg agcaaaattt aagctacaac aaggcaaggc
     5881 ttgaccgaca attgcatgaa gaatctgctt agggttaggc gttttgcgct gcttcgcgat
     5941 gtacgggcca gatatacgcg ttgacattga ttattgacta gttattaata gtaatcaatt
     6001 acggggtcat tagttcatag cccatatatt gagttccgcg ttacataact tacggtaaat
     6061 ggcccgcctg gctgaccgcc caacgacccc cgcccattga cgtcaataat gacgtatgtt
     6121 cccatagtaa cgccaatagg gactttccat tgacgtcaat gggtggagta tttacggtaa
     6181 actgcccact tggcagtaca tcaagtgtat c
//
