LOCUS       sequence-342946-        3210 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  deGFP reporter plasmid with GTAAT PAM upstream of p70a promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3210)
  AUTHORS   Wimmer F, Englert F, Beisel CL
  TITLE     A TXTL-Based Assay to Rapidly Identify PAMs for CRISPR-Cas Systems 
            with Multi-Protein Effector Complexes.
  JOURNAL   Methods Mol Biol. 2022;2433:391-411. doi: 
            10.1007/978-1-0716-1998-8_24.
  PUBMED    34985758
REFERENCE   2  (bases 1 to 3210)
  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: "Methods Mol
            Biol."; date: "2022"; volume: "2433:391-411. doi"; pages: " 
            10.1007/978-1-0716-1998-8_24"
COMMENT     Sequence Label: pGFP-GTAAT
FEATURES             Location/Qualifiers
     source          1..3210
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     151..172
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(367..384)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(538..1126)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(618..637)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1297..2088,2089..2157))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     1920..1939
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2158..2262)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2330..2348
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     RBS             2694..2716
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     primer_bind     complement(2753..2774)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3014..3033)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
ORIGIN
        1 ccgccacaac atcgaggacg gcagcgtgca gctcgccgac cactaccagc agaacacccc
       61 catcggcgac ggccccgtgc tgctgcccga caaccactac ctgagcaccc agtccgccct
      121 gagcaaagac cccaacgaga agcgcgatca catggtcctg ctggagttcg tgaccgccgc
      181 cgggatctaa ctcgagcaaa gcccgccgaa aggcgggctt ttctgtgtcg accgatgccc
      241 ttgagagcct tcaacccagt cagctccttc cggtgggcgc ggggcatgac tatcgtcgcc
      301 gcacttatga ctgtcttctt tatcatgcaa ctcgtaggac aggtgccggc agcgctcttc
      361 cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc
      421 tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat
      481 gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt
      541 ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg
      601 aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc
      661 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt
      721 ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa
      781 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta
      841 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa
      901 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa
      961 ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt
     1021 cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt
     1081 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat
     1141 cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat
     1201 gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc
     1261 aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc
     1321 acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta
     1381 gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga
     1441 cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg
     1501 cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc
     1561 tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat
     1621 cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag
     1681 gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat
     1741 cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa
     1801 ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa
     1861 gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caacacggga
     1921 taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg
     1981 gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc
     2041 acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg
     2101 aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact
     2161 cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat
     2221 atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt
     2281 gccacctgac gtctaagaaa ccattattat catgacatta acctataaaa ataggcgtat
     2341 cacgaggccc tttcgtcttc aagaattctg gcgaatcctt taattaactg accagccaga
     2401 aaacgacctt tctgtggtga aaccggatgc tgcaattcag agcgccagca agtgggggac
     2461 agcagaagac ctgaccgccg cagagtggat gtttgacatg gtgaagacta tcgcaccatc
     2521 agccagaaaa ccgaattttg ctgggtgggc taacgatatc cgcctgatgc gtgaacgtga
     2581 cggacgtaac caccgcgaca tgtgtgtgcg taatcgctgg gcatgctgag ctaacaccgt
     2641 gcgtgttgac aattttacct ctggcggtga taatggttgc agctagcaat aattttgttt
     2701 aactttaaga aggagatata ccatggagct tttcactggc gttgttccca tcctggtcga
     2761 gctggacggc gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggcg agggcgatgc
     2821 cacctacggc aagctgaccc tgaagttcat ctgcaccacc ggcaagctgc ccgtgccctg
     2881 gcccaccctc gtgaccaccc tgacctacgg cgtgcagtgc ttcagccgct accccgacca
     2941 catgaagcag cacgacttct tcaagtccgc catgcccgaa ggctacgtcc aggagcgcac
     3001 catcttcttc aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga
     3061 caccctggtg aaccgcatcg agctgaaggg catcgacttc aaggaggacg gcaacatcct
     3121 ggggcacaag ctggagtaca actacaacag ccacaacgtc tatatcatgg ccgacaagca
     3181 gaagaacggc atcaaggtga acttcaagat
//
