LOCUS       sequence-330475-        4825 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  This vector is designed for efficient cloning of sgRNAs by Golden 
            Gate assembly. The sgRNA insertion leads to replacement of mCherry, 
            resulting in loss of red color of E. coli colony..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4825)
  AUTHORS   Liu X, Kimmey JM, Matarazzo L, de Bakker V, Van Maele L, Sirard JC, 
            Nizet V, Veening JW
  TITLE     Exploration of Bacterial Bottlenecks and Streptococcus pneumoniae 
            Pathogenesis by CRISPRi-Seq.
  JOURNAL   Cell Host Microbe. 2021 Jan 13;29(1):107-120.e6. doi: 
            10.1016/j.chom.2020.10.001. Epub 2020 Oct 28.
  PUBMED    33120116
REFERENCE   2  (bases 1 to 4825)
  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"; doi: 
            "10.1016/j.chom.2020.10.001"; journalName: "Cell Host Microbe"; 
            date: "2021-01-13- 13"; volume: "29"; issue: "1"; pages: 
            "107-120.e6"
COMMENT     Sequence Label: pPEPZ-sgRNAclone
FEATURES             Location/Qualifiers
     source          1..4825
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      38..626
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     527..546
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     misc_feature    2815..2833
                     /label=Tn5 ME
                     /note="hyperactive mosaic end for Tn5 transposase 
                     recognition (Reznikoff et al., 2004)"
     misc_feature    3716..3734
                     /label=Tn5 ME
                     /note="hyperactive mosaic end for Tn5 transposase 
                     recognition (Reznikoff et al., 2004)"
ORIGIN
        1 gttggtacca ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg
       61 taatctgctg cttgcaaaca aaaaaaccac cgctggtagc ggtggtttgt ttgccggatc
      121 aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata
      181 ctgtccttct agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta
      241 catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc
      301 ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg
      361 ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac
      421 agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg
      481 taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt
      541 atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct
      601 cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg
      661 ccttttgctg gccttttgct cacatgttct ttcctgcgtt atcccgcggc cgccaataaa
      721 ttgcttcctt gttttgtcat aattttttta tttaaaaaat tatgacaaaa gtgtgctatt
      781 ctttttatga gaggtgtatg aatatgataa atgtatgtga taaatgtatg tgatgttgga
      841 aaaagaataa aagaacttag aatatcttca aatcttactc aagataagat tgctgaatat
      901 ttgtctttat atttgtcttt gaatcaaagc atgattgcca aaatggaaaa aggtgaaagg
      961 aatatcacga atggatttaa gtaataaagc ttcaaatctt agaaaaaagt tgggagctga
     1021 tggtgaagcg ccgatagata tttttaaatt ggtacaaaag atagaaaatt tgacgctggt
     1081 attttatgga ctcggaaaga atattagcgg agtctgttat aaaggaactc agttcagtct
     1141 cattgcagtc aattcagaca tgtcattagg aaggtaaaga ttttctttag cacatgaact
     1201 gtatcatctt tattatgatg aggtgaagaa gagttcagtc agtcttatct tgattggtga
     1261 aggagatgaa actgaaagaa aagcggatca gcttgcttct tattttttaa ttttcccatc
     1321 ttcactgtat aggatggttg aggaaatcag agaaaatgcc aatagaactc atcttgaagt
     1381 agaagatatt ataaaattgg gtcagtttta tggtatcagt cataaagcta tgttatatag
     1441 attgaggaat gatggatacc ttgatgcaga agaaattaaa aatatggata ttagtgttat
     1501 agagacagct tcaagattag gctatgatac aagtttatat cgtcctttgt cagaaagtaa
     1561 aaaggaaatg gtattaggat aatatattaa ttcaactgaa caacttttag aaaataacag
     1621 aatttcgcaa gggaagtatg aggaactgtt actagatgct ttcagatatg atattgtata
     1681 tgggctagat gaagaggggg gagttgtcgt tgaaaagctt ctagcaaaaa actggacgga
     1741 gtcccgtcca gaaaagtaag cacttgttat tgcaataaaa ttagcctaat tgagagaagt
     1801 ttctatagaa tttttcatat acttaacgag tgctttcacc tttgaatata gtccttccca
     1861 cttatcatca cactctcccc gatagccttt tctagctata tccagtaaag ttacatgctc
     1921 tttaggtaaa agaggtatag cccattcagc agcgacatct ttcgaggtaa tttcaccagt
     1981 agtcactgtt tgccacattc gagctagggt taaaattaca ttacgctcat caccttttat
     2041 cccctcaatt agttctggca aagaatcctt aattgctctt cgaatatctg tcaaaggtac
     2101 ggagacaagt atacttgaag aatcaggacc aaatagagaa atactattct ttcttgcttg
     2161 tgctaaaaca atagccaaat caggatcata gcttggttcc tgaatttgtc cattctcaaa
     2221 ttcacccctg agccactcac cgtatataaa ttctcttttt ggaggatatt gccaagggac
     2281 aacttcactc ctatttataa ccgtaacttc aagtggtcta acagaatccg tatttccaat
     2341 ccttcctgat atagtcatta gtctttctgt tagttttttt cgagttaatt gaggtaaact
     2401 atgattcacg acgactagaa catctacatc gctgttaatg cgtaaaccac catttactgc
     2461 tgaaccaaat agatatactc caactattga acttccaaat aaatctttta cgatttttaa
     2521 tgtttgaatc gcttgatttg gtatttttcc gttaatcaaa ttgctcatga tttcacctcg
     2581 ttgattatgt tcatataccc gaatattttt ttacctacct agtatagcat tttgtgaagt
     2641 ttttttctag tccaagctca caaaaatcca aagtaaccgc tttattaagc cattcttaaa
     2701 taaaaataaa aaaagattaa tagctaaaac tattaatctt atcatatccg caaaaaaccc
     2761 cgcttcggcg gggttttttc gcaattcggt cgacagatct tcgtcggcag cgtcagatgt
     2821 gtataagaga cagccattct acagtttatt cttgacattg cactgtcccc ctggtataat
     2881 aactatatga gacgaggagg aaaattaatg agcaaaggag aagaagataa catggcaatc
     2941 atcaaagaat ttatgcgttt caaagttcac atggaaggtt ctgtaaacgg acacgaattt
     3001 gaaattgaag gtgaaggtga aggccgtcct tatgaaggaa cacaaacggc aaagctgaaa
     3061 gtaacaaaag gcggaccgct tccgtttgca tgggatatcc tttctccgca attcatgtac
     3121 ggttcaaaag catacgtgaa gcatccggct gatattcctg attatttgaa gctgtcattc
     3181 cctgaaggct tcaaatggga gcgtgtgatg aactttgaag atggcggtgt tgttactgtt
     3241 actcaagatt caagccttca agacggtgaa tttatttaca aagtgaagct gcgcggaaca
     3301 aacttcccat ctgacggacc tgtcatgcaa aagaaaacaa tgggctggga agcaagctct
     3361 gaacgcatgt atccagagga cggtgcttta aaaggagaaa tcaaacagcg tttgaagctg
     3421 aaagacggcg gacactatga cgctgaagtg aaaacaactt acaaagcgaa aaagccggtt
     3481 cagcttccag gtgcttacaa cgtaaacatc aaacttgata ttacaagcca caatgaagat
     3541 tatacgattg ttgaacaata tgaacgcgct gaaggccgtc attcaactgg cggaatggat
     3601 gagctttaca aataacgtct cggtttaaga gctatgctgg aaacagcata gcaagtttaa
     3661 ataaggctag tccgttatca acttgaaaaa gtggcaccga gtcggtgctt tttttctgtc
     3721 tcttatacac atctccgagc ccacgagact aaggcgaatc tcgtatgccg tcttctgctt
     3781 gctcgagaaa aaaaaaccgc gcccctgaca gggcgcggtt tttttttcgt aatttattac
     3841 tgaacaagag ggcaatcata tctggaataa tatgcttaaa aagagaagga aaattggtgc
     3901 aaattcattt tcagactatc ttcgtggaag tattcatcaa aatagacaaa aataaatttg
     3961 gataaatcga actcactatt caggaggcat atgagcaatt cgaaaaagaa aagtgtcaaa
     4021 ttgagcctat aggagtagaa gtgaaatagt aagtcctgca tagtggatga gagaaaagtt
     4081 ctccttgaag ttttcctgaa ctatcagtcg catgtcaaac gatatgtagg gtaatgtgag
     4141 aggggatagc gagtagtttt tggttatttt atcaaaaaac ttatatttta ttataccgaa
     4201 tgataaaata taataaaaat gatagaataa ggaaaaaaca tgaatgtcaa aaagataatg
     4261 tcaatttttc aatcctttta tgttgatgtc agtattgagg aactgacttt gactttacca
     4321 atcagttttg taaaaaggtt tgagtatact caaatgactt ttcataagga atcattttta
     4381 ttgattaaag aaaagagaag ggggagtttg agttcatttg ttactcaggc tcgcactatg
     4441 ggtgaaaaag ccaatatgga tgttgttttg gtgttttcga agttatcaga cagtgaaaaa
     4501 aagcaattac ttcaagctag agttccgttt gtagacttta agggaaacct cttcttccct
     4561 ccattgggac tagtactcaa tgcgaatgat actgaagtcc ctaaggaatt aacacctagc
     4621 gaacaattaa cgtggattgc ctttttattg acaaaaggtc aaaaagtagt agatgttgat
     4681 ttgctttcac aagtcactgg acttccaaac tcaacaattt ataggtgttt gaggactttt
     4741 aaagctttat attggttaaa caagcaaaat aagctttaca catatacggt gtcaaagaaa
     4801 gaattattct taaaatccgt gtcat
//
