LOCUS       sequence-225527-        3181 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  expression of gRNA targeting ST6GAL1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3181)
  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 3181)
  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: gh40
FEATURES             Location/Qualifiers
     source          1..3181
                     /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        271..346
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     repeat_region   444..584
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   444..573
                     /label=AAV2 ITR
     rep_origin      659..1114
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(746..765)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     956..977
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(1131..1150)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1250..1272
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1310..1328)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1396..1500
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1501..1569,1570..2361)
                     /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(1719..1738)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2532..3120
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3021..3040
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
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 tgtatcctca agcagcaccc gttttagagc tagaaatagc aagttaaaat
      301 aaggctagtc cgttatcaac ttgaaaaagt ggcaccgagt cggtgctttt ttgttttaga
      361 gctagaaata gcaagttaaa ataaggctag tccgttttta gcgcgtgcgc caattctgca
      421 gacaaatggc tctagagggc cgcaggaacc cctagtgatg gagttggcca ctccctctct
      481 gcgcgctcgc tcgctcactg aggtcgggcg accaaaggtc gcccgacgcc cgggctttgc
      541 ccgggcggcc tcagtgagcg agcgagcgcg cagctgcctg caggggcgcc tgatgcggta
      601 ttttctcctt acgcatctgt gcggtatttc acaccgcata cgtcaaagca accatagtac
      661 gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct
      721 acacttgcca gcgccttagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg
      781 ttcgccggct ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt
      841 gctttacggc acctcgaccc caaaaaactt gatttgggtg atggttcacg tagtgggcca
      901 tcgccctgat agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga
      961 ctcttgttcc aaactggaac aacactcaac tctatctcgg gctattcttt tgatttataa
     1021 gggattttgc cgatttcggt ctattggtta aaaaatgagc tgatttaaca aaaatttaac
     1081 gcgaatttta acaaaatatt aacgtttaca attttatggt gcactctcag tacaatctgc
     1141 tctgatgccg catagttaag ccagccccga cacccgccaa cacccgctga cgcgccctga
     1201 cgggcttgtc tgctcccggc atccgcttac agacaagctg tgaccgtctc cgggagctgc
     1261 atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga gacgaaaggg cctcgtgata
     1321 cgcctatttt tataggttaa tgtcatgata ataatggttt cttagacgtc aggtggcact
     1381 tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg
     1441 tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt
     1501 atgagtattc aacatttccg tgtcgccctt attccctttt ttgcggcatt ttgccttcct
     1561 gtttttgctc acccagaaac gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca
     1621 cgagtgggtt acatcgaact ggatctcaac agcggtaaga tccttgagag ttttcgcccc
     1681 gaagaacgtt ttccaatgat gagcactttt aaagttctgc tatgtggcgc ggtattatcc
     1741 cgtattgacg ccgggcaaga gcaactcggt cgccgcatac actattctca gaatgacttg
     1801 gttgagtact caccagtcac agaaaagcat cttacggatg gcatgacagt aagagaatta
     1861 tgcagtgctg ccataaccat gagtgataac actgcggcca acttacttct gacaacgatc
     1921 ggaggaccga aggagctaac cgcttttttg cacaacatgg gggatcatgt aactcgcctt
     1981 gatcgttggg aaccggagct gaatgaagcc ataccaaacg acgagcgtga caccacgatg
     2041 cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg gcgaactact tactctagct
     2101 tcccggcaac aattaataga ctggatggag gcggataaag ttgcaggacc acttctgcgc
     2161 tcggcccttc cggctggctg gtttattgct gataaatctg gagccggtga gcgtggaagc
     2221 cgcggtatca ttgcagcact ggggccagat ggtaagccct cccgtatcgt agttatctac
     2281 acgacgggga gtcaggcaac tatggatgaa cgaaatagac agatcgctga gataggtgcc
     2341 tcactgatta agcattggta actgtcagac caagtttact catatatact ttagattgat
     2401 ttaaaacttc atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg
     2461 accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc
     2521 aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa
     2581 ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag
     2641 gtaactggct tcagcagagc gcagatacca aatactgttc ttctagtgta gccgtagtta
     2701 ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta
     2761 ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag
     2821 ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg
     2881 gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg
     2941 cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag
     3001 cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc
     3061 cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa
     3121 aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg
     3181 t
//
