LOCUS       sequence-357074-        2989 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  PasCas13b-gRNA targeting EGFP(W58stop) for RNA editing.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2989)
  AUTHORS   Somiya M, Kuroda S
  TITLE     Verification of extracellular vesicle-mediated functional mRNA 
            delivery by RNA editing
  JOURNAL   bioRxiv 2022.01.25.477620
REFERENCE   2  (bases 1 to 2989)
  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: "bioRxiv 
            2022.01.25.477620"
COMMENT     Sequence Label: PasCas13b-gRNA-EGFP(W58stop)
FEATURES             Location/Qualifiers
     source          1..2989
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     150..167
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    284..305
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(320..325,326..343,344..350)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    358..374
                     /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)."
     primer_bind     382..398
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     382..398
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        412..660
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     412..432
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     583..602
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     primer_bind     complement(771..788)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(771..787)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(780..802)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(996..1015)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1115..1137
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1175..1193)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1261..1365
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1366..1434,1435..2226)
                     /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(1584..1603)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2397..2985
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2886..2905
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct
       61 ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag tgagctgata
      121 ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gcggaagagc
      181 gcccaatacg caaaccgcct ctccccgcgc gttggccgat tcattaatgc agctggcacg
      241 acaggtttcc cgactggaaa gcgggcagtg agcgcaacgc aattaatgtg agttagctca
      301 ctcattaggc accccaggct ttacacttta tgcttccggc tcgtatgttg tgtggaattg
      361 tgagcggata acaatttcac gcaggaaaca gctatgacca tgattacgcc agagggccta
      421 tttcccatga ttccttcata tttgcatata cgatacaagg ctgttagaga gataattgga
      481 attaatttga ctgtaaacac aaagatatta gtacaaaata cgtgacgtag aaagtaataa
      541 tttcttgggt agtttgcagt tttaaaatta tgttttaaaa tggactatca tatgcttacc
      601 gtaacttgaa agtatttcga tttcttggct ttatatatct tgtggaaagg acgaaacacc
      661 tggtcacgag ggtgggccag ggcacgggca gcttgccggt ggtgcagatg gttgtggaag
      721 gtccagtttt gaggggctat tacaactttt ttggtaccga gctcgaattc actggccgtc
      781 gttttacaac gtcgtgactg ggaaaaccct ggcgttaccc aacttaatcg ccttgcagca
      841 catccccctt tcgccagctg gcgtaatagc gaagaggccc gcaccgatcg cccttcccaa
      901 cagttgcgca gcctgaatgg cgaatggcgc ctgatgcggt attttctcct tacgcatctg
      961 tgcggtattt cacaccgcat atggtgcact ctcagtacaa tctgctctga tgccgcatag
     1021 ttaagccagc cccgacaccc gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc
     1081 ccggcatccg cttacagaca agctgtgacc gtctccggga gctgcatgtg tcagaggttt
     1141 tcaccgtcat caccgaaacg cgcgagacga aagggcctcg tgatacgcct atttttatag
     1201 gttaatgtca tgataataat ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg
     1261 cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga
     1321 caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat
     1381 ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca
     1441 gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc
     1501 gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca
     1561 atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtat tgacgccggg
     1621 caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca
     1681 gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata
     1741 accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag
     1801 ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg
     1861 gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca
     1921 acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta
     1981 atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct
     2041 ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca
     2101 gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag
     2161 gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat
     2221 tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt
     2281 taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa
     2341 cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga
     2401 gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg
     2461 gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc
     2521 agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag
     2581 aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc
     2641 agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg
     2701 cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac
     2761 accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga
     2821 aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt
     2881 ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag
     2941 cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacg
//
