LOCUS       sequence-68788-e        2286 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Can be used to generate synthetic guide in vitro from T7 promotor.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2286)
  AUTHORS   Waaijers S, Portegijs V, Kerver J, Lemmens BB, Tijsterman M, van den
            Heuvel S, Boxem M
  TITLE     CRISPR/Cas9-Targeted Mutagenesis in Caenorhabditis elegans.
  JOURNAL   Genetics. 2013 Aug 26.
  PUBMED    23979586
REFERENCE   2  (bases 1 to 2286)
  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: "Genetics. 
            2013 Aug 26."
COMMENT     Sequence Label: pMB60
FEATURES             Location/Qualifiers
     source          1..2286
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     2..19
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     3..19
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_RNA        64..139
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        complement(157..175)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(196..212)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(269..286)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(379..967)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(459..478)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(1141..1935)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     promoter        complement(1936..2040)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ctgtaaaacg acggccagtg aattgtaata cgactcacta taagagaccg agagagggtc
       61 tcagttttag agctagaaat agcaagttaa aataaggcta gtccgttatc aacttgaaaa
      121 agtggcaccg agtcggtgct tttaaagctt ttgttccctt tagtgagggt taatttcgag
      181 cttggcgtaa tcatggtcat agctgtttcc tgcagctgca ttaacatggt catagctgtt
      241 tccttgcgta ttgggcgctc tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc
      301 gggtaaagcc tggggtgcct aatgagcaaa aggccagcaa aaggccagga accgtaaaaa
      361 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg
      421 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc
      481 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc
      541 ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc
      601 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg
      661 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc
      721 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga
      781 gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc
      841 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac
      901 caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg
      961 atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc
     1021 acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa
     1081 ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta
     1141 ttagaaaaat tcatccagca gacgataaaa cgcaatacgc tggctatccg gtgccgcaat
     1201 gccatacagc accagaaaac gatccgccca ttcgccgccc agttcttccg caatatcacg
     1261 ggtggccagc gcaatatcct gataacgatc cgccacgccc agacggccgc aatcaataaa
     1321 gccgctaaaa cggccatttt ccaccataat gttcggcagg cacgcatcac catgggtcac
     1381 caccagatct tcgccatccg gcatgctcgc tttcagacgc gcaaacagct ctgccggtgc
     1441 caggccctga tgttcttcat ccagatcatc ctgatccacc aggcccgctt ccatacgggt
     1501 acgcgcacgt tcaatacgat gtttcgcctg atgatcaaac ggacaggtcg ccgggtccag
     1561 ggtatgcaga cgacgcatgg catccgccat aatgctcact ttttctgccg gcgccagatg
     1621 gctagacagc agatcctgac ccggcacttc gcccagcagc agccaatcac ggcccgcttc
     1681 ggtcaccaca tccagcaccg ccgcacacgg aacaccggtg gtggccagcc agctcagacg
     1741 cgccgcttca tcctgcagct cgttcagcgc accgctcaga tcggttttca caaacagcac
     1801 cggacgaccc tgcgcgctca gacgaaacac cgccgcatca gagcagccaa tggtctgctg
     1861 cgcccaatca tagccaaaca gacgttccac ccacgctgcc gggctacccg catgcaggcc
     1921 atcctgttca atcatactct tcctttttca atattattga agcatttatc agggttattg
     1981 tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg
     2041 cacatttccc cgaaaagtgc cacctaaatt gtaagcgtta atattttgtt aaaattcgcg
     2101 ttaaattttt gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg caaaatccct
     2161 tataaatcaa aagaatagac cgagataggg ttgagtggcc gctacagggc gctcccattc
     2221 gccattcagg ctgcgcaact gttgggaagg gcgtttcggt gcgggcctct tcgctattac
     2281 gccagg
//
