LOCUS       sequence-215174-        3025 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Single short guide RNA targeting GTATAGCATACATTATACG.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3025)
  AUTHORS   Rui Y, Varanasi M, Mendes S, Yamagata HM, Wilson DR, Green JJ
  TITLE     Poly(Beta-Amino Ester) Nanoparticles Enable Nonviral Delivery of 
            CRISPR-Cas9 Plasmids for Gene Knockout and Gene Deletion.
  JOURNAL   Mol Ther Nucleic Acids. 2020 Apr 21;20:661-672. doi: 
            10.1016/j.omtn.2020.04.005.
  PUBMED    32380416
REFERENCE   2  (bases 1 to 3025)
  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: "Mol Ther 
            Nucleic Acids."; date: "2020-04-21"; volume: "20:661-672. doi"; 
            pages: " 10.1016/j.omtn.2020.04.005"
COMMENT     Sequence Label: sg1
FEATURES             Location/Qualifiers
     source          1..3025
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(281..1072,1073..1141))
                     /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     904..923
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1142..1246)
                     /gene="bla"
                     /label=AmpR promoter
     promoter        1365..1613
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     1365..1385
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     1536..1555
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     protein_bind    complement(1730..1754)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(1800..1816)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1800..1816)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1813..1835)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1824..1840
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(1848..1854,1855..1872,1873..1878))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1893..1914
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        1973..2168
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      2019..2154
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    2174..2308
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2211..2230)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2265..2284
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(2376..2393)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(join(2547..3025,1..110))
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2627..2646)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg
       61 gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt
      121 tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg
      181 tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta
      241 aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg
      301 aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg
      361 tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc
      421 gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg
      481 agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg
      541 aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag
      601 gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat
      661 caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc
      721 cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc
      781 ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa
      841 ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac
      901 gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt
      961 cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc
     1021 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa
     1081 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca
     1141 tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat
     1201 acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa
     1261 aagtgccacc tggtcgacat tgattattga ctagttgtac aaaaaagcag gctttaaagg
     1321 aaccaattca gtcgactgga tccggtacca aggtcgggca ggaagagggc ctatttccca
     1381 tgattccttc atatttgcat atacgataca aggctgttag agagataatt agaattaatt
     1441 tgactgtaaa cacaaagata ttagtacaaa atacgtgacg tagaaagtaa taatttcttg
     1501 ggtagtttgc agttttaaaa ttatgtttta aaatggacta tcatatgctt accgtaactt
     1561 gaaagtattt cgatttcttg gctttatata tcttgtggaa aggacgaaac accggtatag
     1621 catacattat acggtttcag agctatgctg gaaacagcat agcaagttga aataaggcta
     1681 gtccgttatc aacttgaaaa agtggcaccg agtcggtgct tttttctaga cccagctttc
     1741 ttgtacaaag ttggcattaa agaattctgc ccccgggcgc caagcttggc gtaatcatgg
     1801 tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc
     1861 ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg
     1921 ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagcggat ccgcatctca
     1981 attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca
     2041 gttccgccca ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg
     2101 ccgcctcggc ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct
     2161 tttgcaaaaa gctaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat
     2221 cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact
     2281 catcaatgta tcttatcatg tctggatccg ctgcattaat gaatcggcca acgcgcgggg
     2341 agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg
     2401 gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca
     2461 gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac
     2521 cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac
     2581 aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg
     2641 tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac
     2701 ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc aatgctcacg ctgtaggtat
     2761 ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag
     2821 cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac
     2881 ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt
     2941 gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaaggac agtatttggt
     3001 atctgcgctc tgctgaagcc agtta
//
