LOCUS       sequence-375281-        3027 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses a gRNA for base editing the Kozak sequence of the human 
            NCF1 gene.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3027)
  AUTHORS   Ambrosini C, Destefanis E, Kheir E, Broso F, Alessandrini F, Longhi 
            S, Battisti N, Pesce I, Dassi E, Petris G, Cereseto A, Quattrone A
  TITLE     Translational enhancement by base editing of the Kozak sequence 
            rescues haploinsufficiency.
  JOURNAL   Nucleic Acids Res. 2022 Oct 14;50(18):10756-10771. doi: 
            10.1093/nar/gkac799.
  PUBMED    36165847
REFERENCE   2  (bases 1 to 3027)
  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"; journalName: 
            "Nucleic Acids Res"; date: "2022-10-14- 14"; volume: "50"; issue: 
            "18"; pages: "10756-10771"
COMMENT     Sequence Label: pUC19-sgNCF1
FEATURES             Location/Qualifiers
     source          1..3027
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        86..190
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(191..259,260..1051)
                     /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(409..428)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1222..1810
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1711..1730
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1964..1981
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    2098..2119
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(2134..2139,2140..2157,2158..2164)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2172..2188
                     /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     2177..2199
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     2196..2212
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2196..2212
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        complement(2368..2616)
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     complement(2426..2445)
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     primer_bind     complement(2596..2616)
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     complement(2623..2640)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2623..2639)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2632..2654)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(2848..2867)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2967..2989
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(join(3027,1..18))
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt cttagacgtc
       61 aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca
      121 ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa
      181 aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt ttgcggcatt
      241 ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg ctgaagatca
      301 gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga tccttgagag
      361 ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc tatgtggcgc
      421 ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac actattctca
      481 gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg gcatgacagt
      541 aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca acttacttct
      601 gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg gggatcatgt
      661 aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg acgagcgtga
      721 caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg gcgaactact
      781 tactctagct tcccggcaac aattaataga ctggatggag gcggataaag ttgcaggacc
      841 acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg gagccggtga
      901 gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct cccgtatcgt
      961 agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac agatcgctga
     1021 gataggtgcc tcactgatta agcattggta actgtcagac caagtttact catatatact
     1081 ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga tcctttttga
     1141 taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt
     1201 agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca
     1261 aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct
     1321 ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc ttctagtgta
     1381 gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct
     1441 aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc
     1501 aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca
     1561 gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga
     1621 aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg
     1681 aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt
     1741 cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag
     1801 cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt
     1861 tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgcctt
     1921 tgagtgagct gataccgctc gccgcagccg aacgaccgag cgcagcgagt cagtgagcga
     1981 ggaagcggaa gagcgcccaa tacgcaaacc gcctctcccc gcgcgttggc cgattcatta
     2041 atgcagctgg cacgacaggt ttcccgactg gaaagcgggc agtgagcgca acgcaattaa
     2101 tgtgagttag ctcactcatt aggcacccca ggctttacac tttatgcttc cggctcgtat
     2161 gttgtgtgga attgtgagcg gataacaatt tcacacagga aacagctatg accatgatta
     2221 cgccaagctt gcatgcctgc aggtcgactc tagaaaaaaa gcaccgactc ggtgccactt
     2281 tttcaagttg ataacggact agccttattt aaacttgcta tgctgtttcc agcatagctc
     2341 ttaaacagtc atgggggaca ccttcacggt gtttcgtcct ttccacaaga tatataaagc
     2401 caagaaatcg aaatactttc aagttacggt aagcatatga tagtccattt taaaacataa
     2461 ttttaaaact gcaaactacc caagaaatta ttactttcta cgtcacgtat tttgtactaa
     2521 tatctttgtg tttacagtca aattaattcc aattatctct ctaacagcct tgtatcgtat
     2581 atgcaaatat gaaggaatca tgggaaatag gccctcgaat tcactggccg tcgttttaca
     2641 acgtcgtgac tgggaaaacc ctggcgttac ccaacttaat cgccttgcag cacatccccc
     2701 tttcgccagc tggcgtaata gcgaagaggc ccgcaccgat cgcccttccc aacagttgcg
     2761 cagcctgaat ggcgaatggc gcctgatgcg gtattttctc cttacgcatc tgtgcggtat
     2821 ttcacaccgc atatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca
     2881 gccccgacac ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc
     2941 cgcttacaga caagctgtga ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc
     3001 atcaccgaaa cgcgcgagac gaaaggg
//
