LOCUS       sequence-437045-        2335 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  hNOLC1 atgRNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2335)
  AUTHORS   Fell CW, Schmitt-Ulms C, Tagliaferri DV, Gootenberg JS, Abudayyeh OO
  TITLE     Precise kilobase-scale genomic insertions in mammalian cells using 
            PASTE.
  JOURNAL   Nat Protoc. 2024 Dec 15. doi: 10.1038/s41596-024-01090-z.
  PUBMED    39676077
REFERENCE   2  (bases 1 to 2335)
  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 Protoc.
            2024 Dec 15. doi: 10.1038/s41596-024-01090-z."
COMMENT     Sequence Label: pDY2260 hNOLC1 atgRNA Paired Guide 2
FEATURES             Location/Qualifiers
     source          1..2335
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     393..412
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(631..735)
                     /gene="bla"
                     /label=AmpR promoter
     promoter        842..1090
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     842..862
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     1013..1032
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        1112..1187
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     protein_bind    1192..1229
                     /label=Bxb1 attB
                     /bound_moiety="Mycobacteriophage Bxb1 integrase"
                     /note="Minimal attB site for the Mycopbacteriophage Bxb1 
                     integrase (Voutev and Mann, 2017)"
     rep_origin      complement(1346..1934)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1426..1445)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2105..2335,1..561,562..630))
                     /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"
ORIGIN
        1 ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag
       61 tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat
      121 ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg
      181 caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt
      241 gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag
      301 atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg
      361 accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt
      421 aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct
      481 gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac
      541 tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat
      601 aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat
      661 ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca
      721 aataggggtt ccgcgcacat ttccccgaaa agtgccacct gacgtcgcta gctgtacaaa
      781 aaagcaggct ttaaaggaac caattcagtc gactggatcc ggtaccaagg tcgggcagga
      841 agagggccta tttcccatga ttccttcata tttgcatata cgatacaagg ctgttagaga
      901 gataattaga attaatttga ctgtaaacac aaagatatta gtacaaaata cgtgacgtag
      961 aaagtaataa tttcttgggt agtttgcagt tttaaaatta tgttttaaaa tggactatca
     1021 tatgcttacc gtaacttgaa agtatttcga tttcttggct ttatatatct tgtggaaagg
     1081 acgaaacacc gtattggcca cctctgagag tgttttagag ctagaaatag caagttaaaa
     1141 taaggctagt ccgttatcaa cttgaaaaag tggcaccgag tcggtgcggc cggcttgtcg
     1201 acgacggcgg tctccgtcgt caggatcatc cggctcagag gtttttttta agcttgggcc
     1261 gctcgaggta cctctctaca tatgacatgt gagcaaaagg ccagcaaaag gccaggaacc
     1321 gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca
     1381 aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt
     1441 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
     1501 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc
     1561 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc
     1621 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact
     1681 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg
     1741 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta
     1801 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca
     1861 aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa
     1921 aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg
     1981 aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc
     2041 ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg
     2101 acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat
     2161 ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg
     2221 gccccagtgc tgcaatgata ccgcgagatc cacgctcacc ggctccagat ttatcagcaa
     2281 taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgc
//
