LOCUS       sequence-244226-        2742 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  expression of gRNA targeting LARGE.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2742)
  AUTHORS   Narimatsu Y, Joshi HJ, Zhang Y, Gomes C, Chen YH, Lorenzetti F, 
            Furukawa S, Schjoldager K, Hansen L, Clausen H, Bennett EP, Wandall 
            HH
  TITLE     A validated gRNA library for CRISPR/Cas9 targeting of the human 
            glycosyltransferase genome.
  JOURNAL   Glycobiology. 2018 Jan 5. pii: 4791732. doi: 10.1093/glycob/cwx101.
  PUBMED    29315387
REFERENCE   2  (bases 1 to 2742)
  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: 
            "Glycobiology. 2018 Jan 5. pii: 4791732. doi: 
            10.1093/glycob/cwx101."
COMMENT     Sequence Label: gh144
FEATURES             Location/Qualifiers
     source          1..2742
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..249
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     1..21
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     172..191
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        270..345
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     primer_bind     complement(484..501)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(594..1182)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(674..693)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1353..2144,2145..2213))
                     /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     1976..1995
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2214..2318)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2619..2641
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2633..2650
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2634..2650
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2661..2680
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2661..2679
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 gagggcctat ttcccatgat tccttcatat ttgcatatac gatacaaggc tgttagagag
       61 ataattggaa ttaatttgac tgtaaacaca aagatattag tacaaaatac gtgacgtaga
      121 aagtaataat ttcttgggta gtttgcagtt ttaaaattat gttttaaaat ggactatcat
      181 atgcttaccg taacttgaaa gtatttcgat ttcttggctt tatatatctt gtggaaagga
      241 cgaaacaccg cctggaggtg cgcatgcgcg ttttagagct agaaatagca agttaaaata
      301 aggctagtcc gttatcaact tgaaaaagtg gcaccgagtc ggtgcttttt ttggtacctg
      361 gagcacaaga ctggcctcat gggccttccg ctcactgccc gctttccagt cgggaaacct
      421 gtcgtgccag ctgcattaac atggtcatag ctgtttcctt gcgtattggg cgctctccgc
      481 ttcctcgctc actgactcgc tgcgctcggt cgttcgggta aagcctgggg tgcctaatga
      541 gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat
      601 aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac
      661 ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct
      721 gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg
      781 ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg
      841 ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt
      901 cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg
      961 attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac
     1021 ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga
     1081 aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt
     1141 gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt
     1201 tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga
     1261 ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc
     1321 taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct
     1381 atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata
     1441 actacgatac gggagggctt accatctggc cccagtgctg caatgatacc gcgagaacca
     1501 cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga
     1561 agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga
     1621 gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg
     1681 gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga
     1741 gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt
     1801 gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct
     1861 cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca
     1921 ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat
     1981 accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga
     2041 aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc
     2101 aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg
     2161 caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc
     2221 ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt
     2281 gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca
     2341 cctaaattgt aagcgttaat attttgttaa aattcgcgtt aaatttttgt taaatcagct
     2401 cattttttaa ccaataggcc gaaatcggca aaatccctta taaatcaaaa gaatagaccg
     2461 agatagggtt gagtggccgc tacagggcgc tcccattcgc cattcaggct gcgcaactgt
     2521 tgggaagggc gtttcggtgc gggcctcttc gctattacgc cagctggcga aagggggatg
     2581 tgctgcaagg cgattaagtt gggtaacgcc agggttttcc cagtcacgac gttgtaaaac
     2641 gacggccagt gagcgcgacg taatacgact cactataggg cgaattggcg gaaggccgtc
     2701 aaggccacgt gtcttgtcca gagctcgata tcgaattaat tc
//
