LOCUS       sequence-370606-        4800 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses Marathon-RT pentamutant (D14R-N26R-D74R-N116K-N197R).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4800)
  AUTHORS   Grunewald J, Miller BR, Szalay RN, Cabeceiras PK, Woodilla CJ, Holtz
            EJB, Petri K, Joung JK
  TITLE     Engineered CRISPR prime editors with compact, untethered reverse 
            transcriptases.
  JOURNAL   Nat Biotechnol. 2023 Mar;41(3):337-343. doi: 
            10.1038/s41587-022-01473-1. Epub 2022 Sep 26.
  PUBMED    36163548
REFERENCE   2  (bases 1 to 4800)
  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: 
            "10.1038/s41587-022-01473-1"; journalName: "Nat Biotechnol"; date: 
            "2023-03"; volume: "41"; issue: "3"; pages: "337-343"
COMMENT     Sequence Label: Marathon-RT pentamutant for Split-PE (pEH107)
FEATURES             Location/Qualifiers
     source          1..4800
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        9..388
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        389..592
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     543..563
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     634..653
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        634..652
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             702..722
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2043..2063
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2072..2089
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2112..2129)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2118..2342
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2413..2429)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2413..2429)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2426..2448)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2437..2453
                     /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(2461..2467,2468..2485,2486..2491))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2506..2527
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2644..2661)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2815..3403)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2895..2914)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3574..4365,4366..4434))
                     /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     4197..4216
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4435..4539)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(4618..4637)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 tacgcgttga cattgattat tgactagtta ttaatagtaa tcaattacgg ggtcattagt
       61 tcatagccca tatatggagt tccgcgttac ataacttacg gtaaatggcc cgcctggctg
      121 accgcccaac gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc
      181 aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc
      241 agtacatcaa gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg
      301 gcccgcctgg cattatgccc agtacatgac cttatgggac tttcctactt ggcagtacat
      361 ctacgtatta gtcatcgcta ttaccatggt gatgcggttt tggcagtaca tcaatgggcg
      421 tggatagcgg tttgactcac ggggatttcc aagtctccac cccattgacg tcaatgggag
      481 tttgttttgg caccaaaatc aacgggactt tccaaaatgt cgtaacaact ccgccccatt
      541 gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat ataagcagag ctggtttagt
      601 gaaccgtcag atccgctaga gatccgcggc cgctaatacg actcactata gggagagccg
      661 ccaccatgaa acggacagcc gacggaagcg agttcgagtc accaaagaag aagcggaaag
      721 tcgacaccag caatctgatg gaacagatcc tgagcagccg gaacctgaac cgggcctacc
      781 tgcaggtggt gagacggaaa ggcgctgaag gcgttgatgg catgaagtac accgagctga
      841 aggagcatct ggccaagaac ggcgagacaa tcaagggcca gctgagaacc agaaagtata
      901 agcctcagcc agctagacgg gtggaaatcc ccaagccccg gggcggagtg cggaacctgg
      961 gagtgccaac agtcacagac cggttcatcc agcaggctat cgcccaagtg ctgaccccta
     1021 tctacgagga acagtttcac gaccactctt acggcttccg gcccaagaga tgcgcccagc
     1081 aagccatcct gacagccctg aacatcatga acgatggtaa tgactggatc gtggacatcg
     1141 acctggaaaa gtttttcgat accgtgaatc acgataagct gatgacgctg attggcagaa
     1201 ccatcaagga cggcgacgtg atctctattg tgcgcaagta cctcgtgtcc ggcatcatga
     1261 tcgatgacga gtacgaagat agcatcgtgg gaacacctca gggcggccgg ctgtctcctc
     1321 tgctggccaa catcatgctg aacgagctgg ataaggagat ggaaaaaagg ggcctgaact
     1381 tcgtgcggta cgccgacgac tgcatcatca tggtcggctc cgagatgagc gccaacagag
     1441 tcatgcggaa catcagcaga ttcatcgaag agaagctggg cctgaaagtg aacatgacca
     1501 agtccaaggt ggacagacct agcggactga agtacttggg ctttggcttc tacttcgacc
     1561 ccagagccca ccagttcaag gccaagcctc acgccaagag cgtggctaag ttcaaaaaga
     1621 gaatgaaaga gctgacctgt agaagctggg gcgtgtctaa cagctacaag gtggaaaaac
     1681 tgaatcaact gatcagaggc tggatcaact acttcaagat cggcagcatg aagaccctgt
     1741 gtaaagagct ggacagcaga atcaggtaca gactgcggat gtgcatctgg aagcagtgga
     1801 aaacccctca gaaccaggag aaaaacctgg tcaagcttgg aattgacaga aataccgcca
     1861 gaagagtggc ctatacaggc aagcgaatcg cctacgtgtg caacaagggc gccgtgaacg
     1921 tggctatcag caacaagcgg ctggccagct tcggcctgat ctctatgctg gactactaca
     1981 tcgagaagtg cgtgacctgc tctggcggct caaaaagaac cgccgacggc agcgaattcg
     2041 agcccaagaa gaagaggaaa gtctaaccgg tcatcatcac catcaccatt gagtttaaac
     2101 ccgctgatca gcctcgactg tgccttctag ttgccagcca tctgttgttt gcccctcccc
     2161 cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc ctttcctaat aaaatgagaa
     2221 aattgcatcg cattgtctga gtaggtgtca ttctattctg gggggtgggg tggggcagga
     2281 cagcaagggg gaggattggg aagacaatag caggcatgct ggggatgcgg tgggctctat
     2341 ggcttctgag gcggaaagaa ccagctgggg ctcgataccg tcgacctcta gctagagctt
     2401 ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca
     2461 caacatacga gccggaagca taaagtgtaa agcctagggt gcctaatgag tgagctaact
     2521 cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct
     2581 gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc
     2641 ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca
     2701 ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg
     2761 agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca
     2821 taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa
     2881 cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc
     2941 tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc
     3001 gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct
     3061 gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg
     3121 tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag
     3181 gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta
     3241 cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg
     3301 aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt
     3361 tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt
     3421 ttctacgggg tctgacactc agtggaacga aaactcacgt taagggattt tggtcatgag
     3481 attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat
     3541 ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc
     3601 tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat
     3661 aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc
     3721 acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag
     3781 aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag
     3841 agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt
     3901 ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg
     3961 agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt
     4021 tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc
     4081 tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc
     4141 attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa
     4201 taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg
     4261 aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc
     4321 caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag
     4381 gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt
     4441 cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt
     4501 tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc
     4561 acctgacgtc gacggatcgg gagatcgatc tcccgatccc ctagggtcta ctctcagtac
     4621 aatctgctct gatgccgcat agttaagcca gtatctgctc cctgcttgtg tgttggaggt
     4681 cgctgagtag tgcgcgagca aaatttaagc tacaacaagg caaggcttga ccgacaattg
     4741 catgaagaat ctgcttaggg ttaggcgttt tgcgctgctt cgcgatgtac gggccagata
//
