LOCUS       sequence-221305-        2738 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  expression of gRNA targeting EXT1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2738)
  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 2738)
  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: gh135
FEATURES             Location/Qualifiers
     source          1..2738
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     278..300
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     292..309
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     293..309
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     320..339
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        320..338
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        398..646
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     398..418
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     569..588
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        667..742
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     primer_bind     complement(881..898)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(991..1579)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1071..1090)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1750..2541,2542..2610))
                     /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     2373..2392
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2611..2715)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ctaaattgta agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc
       61 attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga
      121 gatagggttg agtggccgct acagggcgct cccattcgcc attcaggctg cgcaactgtt
      181 gggaagggcg tttcggtgcg ggcctcttcg ctattacgcc agctggcgaa agggggatgt
      241 gctgcaaggc gattaagttg ggtaacgcca gggttttccc agtcacgacg ttgtaaaacg
      301 acggccagtg agcgcgacgt aatacgactc actatagggc gaattggcgg aaggccgtca
      361 aggccacgtg tcttgtccag agctcgatat cgaattcgag ggcctatttc ccatgattcc
      421 ttcatatttg catatacgat acaaggctgt tagagagata attggaatta atttgactgt
      481 aaacacaaag atattagtac aaaatacgtg acgtagaaag taataatttc ttgggtagtt
      541 tgcagtttta aaattatgtt ttaaaatgga ctatcatatg cttaccgtaa cttgaaagta
      601 tttcgatttc ttggctttat atatcttgtg gaaaggacga aacaccgtag aacctggagc
      661 cctcgagttt tagagctaga aatagcaagt taaaataagg ctagtccgtt atcaacttga
      721 aaaagtggca ccgagtcggt gctttttttg gtacctggag cacaagactg gcctcatggg
      781 ccttccgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaacatg
      841 gtcatagctg tttccttgcg tattgggcgc tctccgcttc ctcgctcact gactcgctgc
      901 gctcggtcgt tcgggtaaag cctggggtgc ctaatgagca aaaggccagc aaaaggccag
      961 gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca
     1021 tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca
     1081 ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg
     1141 atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag
     1201 gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt
     1261 tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca
     1321 cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg
     1381 cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt
     1441 tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc
     1501 cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg
     1561 cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg
     1621 gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta
     1681 gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg
     1741 gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg
     1801 ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc
     1861 atctggcccc agtgctgcaa tgataccgcg agaaccacgc tcaccggctc cagatttatc
     1921 agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc
     1981 ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag
     2041 tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat
     2101 ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg
     2161 caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt
     2221 gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag
     2281 atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg
     2341 accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt
     2401 aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct
     2461 gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac
     2521 tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat
     2581 aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat
     2641 ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca
     2701 aataggggtt ccgcgcacat ttccccgaaa agtgccac
//
