LOCUS       sequence-332823-        3318 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  SpyCas9-pegRNA for EMX1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3318)
  AUTHORS   Liu P, Liang SQ, Zheng C, Mintzer E, Zhao YG, Ponnienselvan K, Mir 
            A, Sontheimer EJ, Gao G, Flotte TR, Wolfe SA, Xue W
  TITLE     Improved prime editors enable pathogenic allele correction and 
            cancer modelling in adult mice.
  JOURNAL   Nat Commun. 2021 Apr 9;12(1):2121. doi: 10.1038/s41467-021-22295-w.
  PUBMED    33837189
REFERENCE   2  (bases 1 to 3318)
  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 
            Commun."; date: "2021-04-9"; volume: "12(1):2121. doi"; pages: " 
            10.1038/s41467-021-22295-w"
COMMENT     Sequence Label: Spy EMX1-pegRNA
FEATURES             Location/Qualifiers
     source          1..3318
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    94..115
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(130..135,136..153,154..160)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    168..184
                     /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)."
     primer_bind     173..195
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     192..208
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     192..208
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     227..247
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        229..247
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        294..542
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     294..314
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     465..484
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        563..638
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     primer_bind     complement(691..707)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(692..708)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     complement(732..751)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(733..751)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(761..778)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(761..777)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(770..792)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      919..1374
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1006..1025)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1216..1237
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        1400..1504
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1505..1573,1574..2365)
                     /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     complement(1723..1742)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2536..3124
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3025..3044
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3278..3295
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 caaaccgcct ctccccgcgc gttggccgat tcattaatgc agctggcacg acaggtttcc
       61 cgactggaaa gcgggcagtg agcgcaacgc aattaatgtg agttagctca ctcattaggc
      121 accccaggct ttacacttta tgcttccggc tcgtatgttg tgtggaattg tgagcggata
      181 acaatttcac acaggaaaca gctatgacca tgattacgcc aagcgcgcaa ttaaccctca
      241 ctaaagggaa caaaagctgg agctccaccg cggtggcggc cgcccccttc accgagggcc
      301 tatttcccat gattccttca tatttgcata tacgatacaa ggctgttaga gagataattg
      361 gaattaattt gactgtaaac acaaagatat tagtacaaaa tacgtgacgt agaaagtaat
      421 aatttcttgg gtagtttgca gttttaaaat tatgttttaa aatggactat catatgctta
      481 ccgtaacttg aaagtatttc gatttcttgg ctttatatat cttgtggaaa ggacgaaaca
      541 ccgagtccga gcagaagaag aagttttaga gctagaaata gcaagttaaa ataaggctag
      601 tccgttatca acttgaaaaa gtgggaccga gtcggtcctt gatgtgatgg gagcacttct
      661 tcttctgctc ttttttgata tcaagcttat cgataccgtc gacctcgagg gggggcccgg
      721 tacccaattc gccctatagt gagtcgtatt acgcgcgctc actggccgtc gttttacaac
      781 gtcgtgactg ggaaaaccct ggcgttaccc aacttaatcg ccttgcagca catccccctt
      841 tcgccagctg gcgtaatagc gaagaggccc gcaccgatcg cccttcccaa cagttgcgca
      901 gcctgaatgg cgaatgggac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg
      961 ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct
     1021 tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc
     1081 ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt gattagggtg
     1141 atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt
     1201 ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac cctatctcgg
     1261 tctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta aaaaatgagc
     1321 tgatttaaca aaaatttaac gcgaatttta acaaaatatt aacgcttaca atttaggtgg
     1381 cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa
     1441 tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa
     1501 gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct
     1561 tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg
     1621 tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg
     1681 ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt
     1741 atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga
     1801 cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga
     1861 attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac
     1921 gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg
     1981 ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac
     2041 gatgcctgta gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct
     2101 agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct
     2161 gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg
     2221 gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat
     2281 ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg
     2341 tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata tactttagat
     2401 tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct
     2461 catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa
     2521 gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa
     2581 aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc
     2641 gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag tgtagccgta
     2701 gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct
     2761 gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg
     2821 atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag
     2881 cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc
     2941 cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg
     3001 agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt
     3061 tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg
     3121 gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca
     3181 catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg cctttgagtg
     3241 agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc
     3301 ggaagagcgc ccaatacg
//
