LOCUS       sequence-357449-        4786 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  GsI-IIC RT.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4786)
  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 4786)
  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: GsI-IIC-RT
FEATURES             Location/Qualifiers
     source          1..4786
                     /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"
     regulatory      626..635
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             635..700
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             707..727
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2021..2041
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2050..2067
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2090..2107)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2096..2320
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2391..2407)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2391..2407)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2404..2426)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2415..2431
                     /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(2439..2445,2446..2463,2464..2469))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2484..2505
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2622..2639)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2793..3381)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2873..2892)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3552..4343,4344..4412))
                     /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     4175..4194
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4413..4517)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(4596..4615)
                     /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 gccgcgccac catggactat aaggaccacg acggagacta
      661 caaggatcat gatattgatt acaaagacga tgacgataag atggccccaa agaagaagcg
      721 gaaggtcggt atccacggag tcccagcagc cgcattgttg gaaagaatcc tcgcccgaga
      781 taacctcata actgcactta agcgcgtaga ggctaatcaa ggcgcccctg ggatagatgg
      841 ggtgtcaact gaccaactgc gcgattacat tcgagcacac tggtcaacca tacacgcaca
      901 gcttctcgct gggacatatc gaccagcacc agttcggagg gtcgaaattc caaaaccggg
      961 aggaggcaca aggcagcttg ggatccctac cgttgtcgac cgacttatac agcaggcaat
     1021 tctccaggag ttgacgccta tttttgatcc cgacttctct agctcatcat tcggtttccg
     1081 ccctgggaga aatgcccacg acgctgttcg acaggcacaa ggatacatcc aagaaggata
     1141 tcggtatgtc gtcgatatgg atctcgagaa gtttttcgat agagtgaatc acgatatttt
     1201 gatgagtaga gtagcgcgca aggtgaagga taaaagggtc ctcaagttga ttcgcgcgta
     1261 tctccaagct ggagtcatga tagaaggcgt caaggtgcag acggaagaag ggacaccaca
     1321 aggcggtcct ttgtctcctc ttctcgccaa tatcctcctt gacgacttgg acaaagaact
     1381 ggagaaacga gggctcaagt tttgtcgata cgcggacgac tgtaatattt acgtaaagag
     1441 ccttagggca gggcagcgag tgaagcaaag catccaacgc ttccttgaga aaactctgaa
     1501 actcaaggtt aatgaagaga agtccgcagt cgaccggcca tggaagcgcg cgttcctcgg
     1561 gttctcattt acgcctgaaa ggaaagcgcg gattcgactt gccccacgga gtatccaaag
     1621 attgaagcag aggatacgcc agctcacaaa cccaaattgg tccatatcaa tgccagagcg
     1681 catacacagg gtgaatcagt atgttatggg gtggataggc tactttaggc tcgtcgaaac
     1741 tccaagtgtc ttgcaaacga tagaaggatg gataagacga aggctgaggt tgtgccaatg
     1801 gcttcaatgg aaaagggtac ggactaggat tagggagctt cgcgcccttg gtcttaagga
     1861 aacagccgtg atggaaatag cgaacacccg caaaggcgca tggcgcacta ccaaaacacc
     1921 ccagcttcac caagcattgg gaaagaccta ctggacggcg caaggactta aaagtcttac
     1981 tcaacggtac tttgaactgc ggcaaggtag cgaattcgag cccaagaaga agaggaaagt
     2041 ctaaccggtc atcatcacca tcaccattga gtttaaaccc gctgatcagc ctcgactgtg
     2101 ccttctagtt gccagccatc tgttgtttgc ccctcccccg tgccttcctt gaccctggaa
     2161 ggtgccactc ccactgtcct ttcctaataa aatgagaaaa ttgcatcgca ttgtctgagt
     2221 aggtgtcatt ctattctggg gggtggggtg gggcaggaca gcaaggggga ggattgggaa
     2281 gacaatagca ggcatgctgg ggatgcggtg ggctctatgg cttctgaggc ggaaagaacc
     2341 agctggggct cgataccgtc gacctctagc tagagcttgg cgtaatcatg gtcatagctg
     2401 tttcctgtgt gaaattgtta tccgctcaca attccacaca acatacgagc cggaagcata
     2461 aagtgtaaag cctagggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca
     2521 ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc
     2581 gcggggagag gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg
     2641 cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta
     2701 tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc
     2761 aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag
     2821 catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac
     2881 caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc
     2941 ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt
     3001 aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc
     3061 gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga
     3121 cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta
     3181 ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta
     3241 tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga
     3301 tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg
     3361 cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc tgacactcag
     3421 tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc
     3481 tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact
     3541 tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt
     3601 cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta
     3661 ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta
     3721 tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc
     3781 gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat
     3841 agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt
     3901 atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg
     3961 tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca
     4021 gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta
     4081 agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg
     4141 cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact
     4201 ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg
     4261 ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt
     4321 actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga
     4381 ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc
     4441 atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa
     4501 caaatagggg ttccgcgcac atttccccga aaagtgccac ctgacgtcga cggatcggga
     4561 gatcgatctc ccgatcccct agggtctact ctcagtacaa tctgctctga tgccgcatag
     4621 ttaagccagt atctgctccc tgcttgtgtg ttggaggtcg ctgagtagtg cgcgagcaaa
     4681 atttaagcta caacaaggca aggcttgacc gacaattgca tgaagaatct gcttagggtt
     4741 aggcgttttg cgctgcttcg cgatgtacgg gccagatata cgcgtt
//
