LOCUS       sequence-357448-        5985 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  MCP-M-MLV-RT (MS2-N55K-MMLVRT).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5985)
  AUTHORS   Liu B, Dong X, Cheng H, Zheng C, Chen Z, Rodriguez TC, Liang SQ, Xue
            W, Sontheimer EJ
  TITLE     A split prime editor with untethered reverse transcriptase and 
            circular RNA template.
  JOURNAL   Nat Biotechnol. 2022 Apr 4. pii: 10.1038/s41587-022-01255-9. doi: 
            10.1038/s41587-022-01255-9.
  PUBMED    35379962
REFERENCE   2  (bases 1 to 5985)
  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. 2022 Apr 4. pii: 10.1038/s41587-022-01255-9. doi: 
            10.1038/s41587-022-01255-9."
COMMENT     Sequence Label: MCP-M-MLV-RT
FEATURES             Location/Qualifiers
     source          1..5985
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     626..645
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        626..644
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      652..661
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             661..1047
                     /codon_start=1
                     /product="bacteriophage MS2 coat protein"
                     /label=MS2-N55K
                     /note="binds with high affinity to a specific stem-loop 
                     structure in the viral RNA (Lim et al., 1994)"
                     /translation="ASNFTQFVLVDNGGTGDVTVAPSNFANGVAEWISSNSRSQAYKVT
                     CSVRQSSAQKRKYTIKVEVPKVATQTVGGVELPVAAWRSYLNMELTIPIFATNSDCELI
                     VKAMQGLLKDGNPIPSAIAANSGIY"
     CDS             3220..3240
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3249..3266
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(3289..3306)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3295..3519
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(3590..3606)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3590..3606)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3603..3625)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3614..3630
                     /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(3638..3644,3645..3662,3663..3668))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3683..3704
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3821..3838)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3992..4580)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4072..4091)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4751..5542,5543..5611))
                     /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     5374..5393
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5612..5716)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(5795..5814)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctggttta gtgaaccgtc
      601 agatccgcta gagatccgcg gccgctaata cgactcacta tagggagagc cgccaccatg
      661 gcttcaaact ttactcagtt cgtgctcgtg gacaatggtg ggacagggga tgtgacagtg
      721 gctccttcta atttcgctaa tggggtggca gagtggatca gctccaactc acggagccag
      781 gcctacaagg tgacatgcag cgtcaggcag tctagtgccc agaagagaaa gtataccatc
      841 aaggtggagg tccccaaagt ggctacccag acagtgggcg gagtcgaact gcctgtcgcc
      901 gcttggaggt cctacctgaa catggagctc actatcccaa ttttcgctac caattctgac
      961 tgtgaactca tcgtgaaggc aatgcagggg ctcctcaaag acggtaatcc tatcccttcc
     1021 gccatcgccg ctaactcagg tatctactct ggaggatcta gcggaggatc ctctggcagc
     1081 gagacaccag gaacaagcga gtcagcaaca ccagagagca gtggcggcag cagcggcggc
     1141 agcagcaccc taaatataga agatgagtat cggctacatg agacctcaaa agagccagat
     1201 gtttctctag ggtccacatg gctgtctgat tttcctcagg cctgggcgga aaccgggggc
     1261 atgggactgg cagttcgcca agctcctctg atcatacctc tgaaagcaac ctctaccccc
     1321 gtgtccataa aacaataccc catgtcacaa gaagccagac tggggatcaa gccccacata
     1381 cagagactgt tggaccaggg aatactggta ccctgccagt ccccctggaa cacgcccctg
     1441 ctacccgtta agaaaccagg gactaatgat tataggcctg tccaggatct gagagaagtc
     1501 aacaagcggg tggaagacat ccaccccacc gtgcccaacc cttacaacct cttgagcggg
     1561 ctcccaccgt cccaccagtg gtacactgtg cttgatttaa aggatgcctt tttctgcctg
     1621 agactccacc ccaccagtca gcctctcttc gcctttgagt ggagagatcc agagatggga
     1681 atctcaggac aattgacctg gaccagactc ccacagggtt tcaaaaacag tcccaccctg
     1741 tttaatgagg cactgcacag agacctagca gacttccgga tccagcaccc agacttgatc
     1801 ctgctacagt acgtggatga cttactgctg gccgccactt ctgagctaga ctgccaacaa
     1861 ggtactcggg ccctgttaca aaccctaggg aacctcgggt atcgggcctc ggccaagaaa
     1921 gcccaaattt gccagaaaca ggtcaagtat ctggggtatc ttctaaaaga gggtcagaga
     1981 tggctgactg aggccagaaa agagactgtg atggggcagc ctactccgaa gacccctcga
     2041 caactaaggg agttcctagg gaaggcaggc ttctgtcgcc tcttcatccc tgggtttgca
     2101 gaaatggcag cccccctgta ccctctcacc aaaccgggga ctctgtttaa ttggggccca
     2161 gaccaacaaa aggcctatca agaaatcaag caagctcttc taactgcccc agccctgggg
     2221 ttgccagatt tgactaagcc ctttgaactc tttgtcgacg agaagcaggg ctacgccaaa
     2281 ggtgtcctaa cgcaaaaact gggaccttgg cgtcggccgg tggcctacct gtccaaaaag
     2341 ctagacccag tagcagctgg gtggccccct tgcctacgga tggtagcagc cattgccgta
     2401 ctgacaaagg atgcaggcaa gctaaccatg ggacagccac tagtcattct ggccccccat
     2461 gcagtagagg cactagtcaa acaacccccc gaccgctggc tttccaacgc ccggatgact
     2521 cactatcagg ccttgctttt ggacacggac cgggtccagt tcggaccggt ggtagccctg
     2581 aacccggcta cgctgctccc actgcctgag gaagggctgc aacacaactg ccttgatatc
     2641 ctggccgaag cccacggaac ccgacccgac ctaacggacc agccgctccc agacgccgac
     2701 cacacctggt acacggatgg aagcagtctc ttacaagagg gacagcgtaa ggcgggagct
     2761 gcggtgacca ccgagaccga ggtaatctgg gctaaagccc tgccagccgg gacatccgct
     2821 cagcgggctg aactgatagc actcacccag gccctaaaga tggcagaagg taagaagcta
     2881 aatgtttata ctgatagccg ttatgctttt gctactgccc atatccatgg agaaatatac
     2941 agaaggcgtg ggtggctcac atcagaaggc aaagagatca aaaataaaga cgagatcttg
     3001 gccctactaa aagccctctt tctgcccaaa agacttagca taatccattg tccaggacat
     3061 caaaagggac acagcgccga ggctagaggc aaccggatgg ctgaccaagc ggcccgaaag
     3121 gcagccatca cagagactcc agacacctct accctcctca tagaaaattc atcaccctct
     3181 ggcggctcaa aaagaaccgc cgacggcagc gaattcgagc ccaagaagaa gaggaaagtc
     3241 taaccggtca tcatcaccat caccattgag tttaaacccg ctgatcagcc tcgactgtgc
     3301 cttctagttg ccagccatct gttgtttgcc cctcccccgt gccttccttg accctggaag
     3361 gtgccactcc cactgtcctt tcctaataaa atgagaaaat tgcatcgcat tgtctgagta
     3421 ggtgtcattc tattctgggg ggtggggtgg ggcaggacag caagggggag gattgggaag
     3481 acaatagcag gcatgctggg gatgcggtgg gctctatggc ttctgaggcg gaaagaacca
     3541 gctggggctc gataccgtcg acctctagct agagcttggc gtaatcatgg tcatagctgt
     3601 ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa
     3661 agtgtaaagc ctagggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac
     3721 tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg
     3781 cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc
     3841 gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat
     3901 ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca
     3961 ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc
     4021 atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc
     4081 aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg
     4141 gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta
     4201 ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg
     4261 ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac
     4321 acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag
     4381 gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat
     4441 ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat
     4501 ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc
     4561 gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct gacactcagt
     4621 ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct
     4681 agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt
     4741 ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc
     4801 gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac
     4861 catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat
     4921 cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg
     4981 cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata
     5041 gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta
     5101 tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt
     5161 gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag
     5221 tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa
     5281 gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc
     5341 gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt
     5401 taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc
     5461 tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta
     5521 ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa
     5581 taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca
     5641 tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac
     5701 aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgacgtcgac ggatcgggag
     5761 atcgatctcc cgatccccta gggtctactc tcagtacaat ctgctctgat gccgcatagt
     5821 taagccagta tctgctccct gcttgtgtgt tggaggtcgc tgagtagtgc gcgagcaaaa
     5881 tttaagctac aacaaggcaa ggcttgaccg acaattgcat gaagaatctg cttagggtta
     5941 ggcgttttgc gctgcttcgc gatgtacggg ccagatatac gcgtt
//
