LOCUS       sequence-332824-        3296 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  SauCas9 nicking sgRNA for EMX1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3296)
  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 3296)
  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: SauCas9 EMX1-nicking sgRNA
FEATURES             Location/Qualifiers
     source          1..3296
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     112..131
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     365..382
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    499..520
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(535..540,541..558,559..565)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    573..589
                     /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     578..600
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     597..613
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     597..613
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     632..652
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        634..652
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        699..947
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     699..719
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     870..889
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        971..1046
                     /label=Sa gRNA scaffold
                     /note="guide RNA scaffold for the Staphylococcus aureus 
                     CRISPR/Cas9 system"
     primer_bind     complement(1074..1090)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1075..1091)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     complement(1115..1134)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1116..1134)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1144..1161)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1144..1160)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1153..1175)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      1302..1757
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1389..1408)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1599..1620
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        1783..1887
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1888..1956,1957..2748)
                     /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(2106..2125)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      join(2919..3296,1..211)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta
       61 tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc
      121 ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg
      181 atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt
      241 cctggccttt tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt
      301 ggataaccgt attaccgcct ttgagtgagc tgataccgct cgccgcagcc gaacgaccga
      361 gcgcagcgag tcagtgagcg aggaagcgga agagcgccca atacgcaaac cgcctctccc
      421 cgcgcgttgg ccgattcatt aatgcagctg gcacgacagg tttcccgact ggaaagcggg
      481 cagtgagcgc aacgcaatta atgtgagtta gctcactcat taggcacccc aggctttaca
      541 ctttatgctt ccggctcgta tgttgtgtgg aattgtgagc ggataacaat ttcacacagg
      601 aaacagctat gaccatgatt acgccaagcg cgcaattaac cctcactaaa gggaacaaaa
      661 gctggagctc caccgcggtg gcggccgccc ccttcaccga gggcctattt cccatgattc
      721 cttcatattt gcatatacga tacaaggctg ttagagagat aattggaatt aatttgactg
      781 taaacacaaa gatattagta caaaatacgt gacgtagaaa gtaataattt cttgggtagt
      841 ttgcagtttt aaaattatgt tttaaaatgg actatcatat gcttaccgta acttgaaagt
      901 atttcgattt cttggcttta tatatcttgt ggaaaggacg aaacaccgca gcaagcagca
      961 ctctgccctc gttttagtac tctggaaaca gaatctacta aaacaaggca aaatgccgtg
     1021 tttatctcgt caacttgttg gcgagatttt tttgaattcg atatcaagct tatcgatacc
     1081 gtcgacctcg agggggggcc cggtacccaa ttcgccctat agtgagtcgt attacgcgcg
     1141 ctcactggcc gtcgttttac aacgtcgtga ctgggaaaac cctggcgtta cccaacttaa
     1201 tcgccttgca gcacatcccc ctttcgccag ctggcgtaat agcgaagagg cccgcaccga
     1261 tcgcccttcc caacagttgc gcagcctgaa tggcgaatgg gacgcgccct gtagcggcgc
     1321 attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct
     1381 agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg
     1441 tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga
     1501 ccccaaaaaa cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt
     1561 ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg
     1621 aacaacactc aaccctatct cggtctattc ttttgattta taagggattt tgccgatttc
     1681 ggcctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt ttaacaaaat
     1741 attaacgctt acaatttagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt
     1801 ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg
     1861 cttcaataat attgaaaaag gaagagtatg agtattcaac atttccgtgt cgcccttatt
     1921 cccttttttg cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta
     1981 aaagatgctg aagatcagtt gggtgcacga gtgggttaca tcgaactgga tctcaacagc
     2041 ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc caatgatgag cacttttaaa
     2101 gttctgctat gtggcgcggt attatcccgt attgacgccg ggcaagagca actcggtcgc
     2161 cgcatacact attctcagaa tgacttggtt gagtactcac cagtcacaga aaagcatctt
     2221 acggatggca tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact
     2281 gcggccaact tacttctgac aacgatcgga ggaccgaagg agctaaccgc ttttttgcac
     2341 aacatggggg atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata
     2401 ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta
     2461 ttaactggcg aactacttac tctagcttcc cggcaacaat taatagactg gatggaggcg
     2521 gataaagttg caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat
     2581 aaatctggag ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt
     2641 aagccctccc gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga
     2701 aatagacaga tcgctgagat aggtgcctca ctgattaagc attggtaact gtcagaccaa
     2761 gtttactcat atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag
     2821 gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac
     2881 tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc
     2941 gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat
     3001 caagagctac caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat
     3061 actgttcttc tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct
     3121 acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt
     3181 cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg
     3241 gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact gagata
//
