LOCUS       sequence-415835-        5313 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Recruited B103 DNA polymerase with a C-terminal N5 coiled coil (CC) 
            domain, expressed from CMV or T7 promoters. .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5313)
  AUTHORS   Ferreira da Silva J, Tou CJ, King EM, Eller ML, Rufino-Ramos D, Ma 
            L, Cromwell CR, Metovic J, Benning FMC, Chao LH, Eichler FS, 
            Kleinstiver BP
  TITLE     Click editing enables programmable genome writing using DNA 
            polymerases and HUH endonucleases.
  JOURNAL   Nat Biotechnol. 2024 Jul 22. doi: 10.1038/s41587-024-02324-x.
  PUBMED    39039307
REFERENCE   2  (bases 1 to 5313)
  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: "Nat 
            Biotechnol. 2024 Jul 22. doi: 10.1038/s41587-024-02324-x."
COMMENT     Sequence Label: Recruited polymerase expression -
            pCMV-T7-B103...
FEATURES             Location/Qualifiers
     source          1..5313
                     /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             2182..2202
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2328..2345
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2368..2385)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2374..2598
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2669..2685)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2669..2685)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2682..2704)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2693..2709
                     /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(2717..2723,2724..2741,2742..2747))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2762..2783
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2900..2917)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3071..3659)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3151..3170)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3830..4621,4622..4690))
                     /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     4453..4472
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4691..4795)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(4874..4893)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        join(5065..5313,1..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 atagggagag ccgccaccat gcctagaaag
      421 atgttcagct gcgatttcga aaccacaacc aagctggacg actgcagagt gtgggcttac
      481 ggctacatgg aaatcggcaa cctggataac tacaagattg gaaacagcct ggatgagttc
      541 atgcagtggg tgatggaaat ccaggccgac ctgtacttcc acaacctgaa gtttgacggc
      601 gccttcatag tgaactggct ggaaagacac ggcttcaagt ggagcaacga gggcctgccc
      661 aacacctaca ataccatcat ttctaagatg ggccagtggt acatgatcga tatctgtttc
      721 ggctataagg gcaagcggaa gctgcacacc gtgatctacg acagcctgaa gaagctgcca
      781 ttccctgtca aaaagatcgc caaagatttt cagctgcctc tgctgaaagg cgatatcgac
      841 taccacaagg agcggcctgt cggccacgaa atcacaccag aggaatatga atacatcaag
      901 aatgacatcg agatcatcgc ccgggccctg gacatccagt tcaagcaagg cctggataga
      961 atgaccgctg gctctgattc tctgaagggg ttcaaagata tcctgagcac caagaagttc
     1021 aacaaggtgt tccccaagtt aagcctgcct atggacaagg aaatccgcta cgcctatagg
     1081 ggcggcttta catggctgaa cgacaaatac aaggaaaagg agattggaga aggcatggtg
     1141 ttcgatgtga actctctgta tccttcccag atgtacagca gacctctgcc ttacggcgcc
     1201 cccatcgtgt tccagggcaa gtacgagaag gacgagcagt accccctgta catccaaaga
     1261 atcagattcg agttcgagct gaaagagggc tatatcccca ccatccagat caaaaaaaac
     1321 cccttcttca agggcaatga gtacctgaag aactccggcg gagagcctgt tgaactgtac
     1381 ctgaccaacg tggatctgga actgatccag gagcactacg agctgtacaa cgtggagtac
     1441 attgacggct ttaaatttag agaaaaaacc ggcttgttta aggacttcat cgacaagtgg
     1501 acctacgtga agacacacga ggaaggagca aagaaacagc tggccaagct gatgctgaac
     1561 agcctgtacg gcaagtttgc cagcaatcct gacgtgaccg gcaaagttcc atacctcaaa
     1621 gaggacggat ctctgggctt ccgggtgggc gacgaagaat acaaggaccc cgtgtacacc
     1681 cctatgggag tgtttatcac agcctgggcc agattcacca ccatcaccgc cgcccaggct
     1741 tgttacgacc ggatcatcta ctgcgacaca gacagcatcc acctgacagg caccgaggtg
     1801 cctgagatca tcaaggacat cgtggaccct aagaagctgg gctactgggc tcatgagtcc
     1861 accttcaaaa gagccaagta cctgagacag aagacctaca tccaggacat ctacatgaag
     1921 gaagttgacg gaaagctggt ggagtgcagc cctgatgagg ccaccacaac caagttctcc
     1981 gtgaagtgcg ccggcatgac agataccatc aaaaagaagg tgacattcga caacttcaag
     2041 gtgggcttca gcagcaaggg aaagcctaaa ccagtgcaag tgaacggcgg agtggtgctg
     2101 gtagacagcg tgttcacaat caagtccggc ggaagctctg gtggcagcaa gcggaccgcc
     2161 gacggctctg aattcgagag ccctaagaag aaaagaaagg tgagcggtgg gagcagcggc
     2221 gggtcaggcg agatcgccgc actggaagcc aagatcgctg ccctgaaggc aaagaacgcc
     2281 gctctgaagg ccgagatcgc cgccctggaa gccggatact gaccggtcat catcaccatc
     2341 accattgagt ttaaacccgc tgatcagcct cgactgtgcc ttctagttgc cagccatctg
     2401 ttgtttgccc ctcccccgtg ccttccttga ccctggaagg tgccactccc actgtccttt
     2461 cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct attctggggg
     2521 gtggggtggg gcaggacagc aagggggagg attgggaaga caatagcagg catgctgggg
     2581 atgcggtggg ctctatggct tctgaggcgg aaagaaccag ctggggctcg ataccgtcga
     2641 cctctagcta gagcttggcg taatcatggt catagctgtt tcctgtgtga aattgttatc
     2701 cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc tagggtgcct
     2761 aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa
     2821 acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta
     2881 ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc
     2941 gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg
     3001 caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
     3061 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
     3121 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
     3181 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
     3241 cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg
     3301 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
     3361 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
     3421 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
     3481 agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga
     3541 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
     3601 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
     3661 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag
     3721 ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat
     3781 gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct
     3841 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac
     3901 tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa
     3961 tgataccgcg cgaaccacgc tcaccggctc cagatttatc agcaataaac cagccagccg
     4021 gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt
     4081 gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca
     4141 ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt
     4201 cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct
     4261 tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg
     4321 cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg
     4381 agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg
     4441 cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa
     4501 aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt
     4561 aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt
     4621 gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt
     4681 gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca
     4741 tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat
     4801 ttccccgaaa agtgccacct gacgtcgacg gatcgggaga tcgatctccc gatcccctag
     4861 ggtcgactct cagtacaatc tgctctgatg ccgcatagtt aagccagtat ctgctccctg
     4921 cttgtgtgtt ggaggtcgct gagtagtgcg cgagcaaaat ttaagctaca acaaggcaag
     4981 gcttgaccga caattgcatg aagaatctgc ttagggttag gcgttttgcg ctgcttcgcg
     5041 atgtacgggc cagatatacg cgttgacatt gattattgac tagttattaa tagtaatcaa
     5101 ttacggggtc attagttcat agcccatata tggagttccg cgttacataa cttacggtaa
     5161 atggcccgcc tggctgaccg cccaacgacc cccgcccatt gacgtcaata atgacgtatg
     5221 ttcccatagt aacgccaata gggactttcc attgacgtca atgggtggag tatttacggt
     5281 aaactgccca cttggcagta catcaagtgt atc
//
