LOCUS       sequence-346344-        3435 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Human expression plasmid for SpCas9 sgRNA with blasticidin 
            co-selection.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3435)
  TITLE     Prime editing plasmids
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 3435)
  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: 
            "Unpublished"
COMMENT     Sequence Label: BPK1520_blastR
FEATURES             Location/Qualifiers
     source          1..3435
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        41..420
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        421..624
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     575..595
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      643..652
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             649..1047
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAMPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     polyA_signal    1073..1194
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1110..1129)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1164..1183
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        1237..1485
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     1237..1257
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     1408..1427
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        1507..1582
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     rep_origin      complement(1686..2274)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1766..1785)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2445..3236,3237..3305))
                     /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     3068..3087
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3306..3410)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacgtcgcta gctctagagg gcctgaattc tcgaggcgtt gacattgatt attgactagt
       61 tattaatagt aatcaattac ggggtcatta gttcatagcc catatatgga gttccgcgtt
      121 acataactta cggtaaatgg cccgcctggc tgaccgccca acgacccccg cccattgacg
      181 tcaataatga cgtatgttcc catagtaacg ccaataggga ctttccattg acgtcaatgg
      241 gtggagtatt tacggtaaac tgcccacttg gcagtacatc aagtgtatca tatgccaagt
      301 acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc ccagtacatg
      361 accttatggg actttcctac ttggcagtac atctacgtat tagtcatcgc tattaccatg
      421 gtgatgcggt tttggcagta catcaatggg cgtggatagc ggtttgactc acggggattt
      481 ccaagtctcc accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac
      541 tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg
      601 tgggaggtct atataagcag agctctctgg ctaactaccg gtgccaccat ggccatgcct
      661 ttgtctcaag aagaatccac cctcattgaa agagcaacgg ctacaatcaa cagcatcccc
      721 atctctgaag actacagcgt cgccagcgca gctctctcta gcgacggccg catcttcact
      781 ggtgtcaatg tatatcattt tactggggga ccttgtgcag aactcgtggt gctgggcact
      841 gctgctgctg cggcagctgg caacctgact tgtatcgtcg cgatcggaaa tgagaacagg
      901 ggcatcttga gcccctgcgg acggtgccga caggtgcttc tcgatctgca tcctgggatc
      961 aaagccatag tgaaggacag tgatggacag ccgacggcag ttgggattcg tgaattgctg
     1021 ccctctggtt atgtgtggga gggctgagaa ttcatcgata ccgtcgagat ctaacttgtt
     1081 tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc
     1141 atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt
     1201 ctggaaattc gatccggtac caaggtcggg caggaagagg gcctatttcc catgattcct
     1261 tcatatttgc atatacgata caaggctgtt agagagataa ttagaattaa tttgactgta
     1321 aacacaaaga tattagtaca aaatacgtga cgtagaaagt aataatttct tgggtagttt
     1381 gcagttttaa aattatgttt taaaatggac tatcatatgc ttaccgtaac ttgaaagtat
     1441 ttcgatttct tggctttata tatcttgtgg aaaggacgaa acaccggaga cgattaatgc
     1501 gtctccgttt tagagctaga aatagcaagt taaaataagg ctagtccgtt atcaacttga
     1561 aaaagtggca ccgagtcggt gcttttttta agcttgggcc gctcgaggta cctctctaca
     1621 tatgacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg
     1681 gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag
     1741 aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc
     1801 gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg
     1861 ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt
     1921 cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc
     1981 ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc
     2041 actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg
     2101 tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca
     2161 gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc
     2221 ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat
     2281 cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt
     2341 ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt
     2401 tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc
     2461 agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc
     2521 gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata
     2581 ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg
     2641 gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc
     2701 cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct
     2761 acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa
     2821 cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt
     2881 cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca
     2941 ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac
     3001 tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca
     3061 atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt
     3121 tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc
     3181 actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca
     3241 aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata
     3301 ctcatactct tcctttttca atattattga agcatttatc agggttattg tctcatgagc
     3361 ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc
     3421 cgaaaagtgc cacct
//
