LOCUS       sequence-440913-        2296 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Golden gate backbone for mammalian expression of Sp-gRNA with 
            3'-com.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2296)
  AUTHORS   Cowan QT, Gu S, Gu W, Ranzau BL, Simonson TS, Komor AC
  TITLE     Development of multiplexed orthogonal base editor (MOBE) systems.
  JOURNAL   Nat Biotechnol. 2024 May 21. doi: 10.1038/s41587-024-02240-0.
  PUBMED    38773305
REFERENCE   2  (bases 1 to 2296)
  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 
            Biotechnol. 2024 May 21. doi: 10.1038/s41587-024-02240-0."
COMMENT     Sequence Label: 3'-com-gRNA GG backbone (MOBE1-4)
FEATURES             Location/Qualifiers
     source          1..2296
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     65..84
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        161..236
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     rep_origin      complement(359..947)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(439..458)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1118..1909,1910..1978))
                     /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     1741..1760
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1979..2083)
                     /gene="bla"
                     /label=AmpR promoter
     promoter        join(2190..2296,1..142)
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     2190..2210
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
ORIGIN
        1 tacgtgacgt agaaagtaat aatttcttgg gtagtttgca gttttaaaat tatgttttaa
       61 aatggactat catatgctta ccgtaacttg aaagtatttc gatttcttgg ctttatatat
      121 cttgtggaaa ggacgaaaca ccgggtcttc gagaagacct gttttagagc tagaaatagc
      181 aagttaaaat aaggctagtc cgttatcaac ttgaaaaagt ggcaccgagt cggtgcctga
      241 atgcctgcga gcatcttttt ttaagcttgg gccgctcgag gtacctctct acatatgaca
      301 tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt
      361 tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc
      421 gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct
      481 ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg
      541 tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca
      601 agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact
      661 atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta
      721 acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta
      781 actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct
      841 tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt
      901 tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga
      961 tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca
     1021 tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat
     1081 caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg
     1141 cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt
     1201 agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag
     1261 acccacgctc accggctcca gatttatcag caataaacca gccagccgga agggccgagc
     1321 gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag
     1381 ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca
     1441 tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa
     1501 ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga
     1561 tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata
     1621 attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca
     1681 agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg
     1741 ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg
     1801 ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg
     1861 cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag
     1921 gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac
     1981 tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca
     2041 tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag
     2101 tgccacctga cgtcgctagc tgtacaaaaa agcaggcttt aaaggaacca attcagtcga
     2161 ctggatccgg taccaaggtc gggcaggaag agggcctatt tcccatgatt ccttcatatt
     2221 tgcatatacg atacaaggct gttagagaga taattagaat taatttgact gtaaacacaa
     2281 agatattagt acaaaa
//
