LOCUS       sequence-229420-        2183 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Encodes sgRNA targeting position 3 in pColE1_70a_deGFP_KanR.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2183)
  AUTHORS   Collias D, Marshall R, Collins SP, Beisel CL, Noireaux V
  TITLE     An educational module to explore CRISPR technologies with a 
            cell-free transcription-translation system.
  JOURNAL   Synth Biol (Oxf). 2019 Jan 21;4(1):ysz005. doi: 
            10.1093/synbio/ysz005. eCollection 2019.
  PUBMED    32995532
REFERENCE   2  (bases 1 to 2183)
  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: "Synth Biol 
            (Oxf)."; date: "2019-01-21"; volume: "4(1):ysz005. doi"; pages: " 
            10.1093/synbio/ysz005. eCollection 2019"
COMMENT     Sequence Label: pColE1_sgRNA_pos3_AmpR
FEATURES             Location/Qualifiers
     source          1..2183
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        39..73
                     /label=J23119 promoter
                     /note="bacterial promoter (Registry of Standard Biological 
                     Parts BBa_J23119)"
     misc_RNA        94..169
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     CDS             complement(join(431..1222,1223..1291))
                     /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     1054..1073
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1292..1396)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      1499..2087
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1988..2007
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cgtgcggccc gagagcatcg atcctagcat ggatgcattt gacagctagc tcagtcctag
       61 gtataatgct agcaattttg tttaacttta agagttttag agctagaaat agcaagttaa
      121 aataaggcta gtccgttatc aacttgaaaa agtggcaccg agtcggtgct ttttttccgg
      181 cttatcggtc agtttcacct gatttacgta aaaacccgct tcggcgggtt tttgcttttg
      241 gaggggcaga aagatgaatg actgtccacg acgctatacc caaaagaaaa agcttgaaag
      301 aatcgatcga tcccgaatga gtagtacagc ggccgcgatt atcaaaaagg atcttcacct
      361 agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt
      421 ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc
      481 gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac
      541 catctggccc cagtgctgca atgataccgc gggacccacg ctcaccggct ccagatttat
      601 cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg
      661 cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata
      721 gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta
      781 tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt
      841 gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag
      901 tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa
      961 gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc
     1021 gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt
     1081 taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc
     1141 tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta
     1201 ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa
     1261 taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca
     1321 tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac
     1381 aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgtcatgacc aaaatccctt
     1441 aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt
     1501 gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag
     1561 cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta actggcttca
     1621 gcagagcgca gataccaaat actgttcttc tagtgtagcc gtagttaggc caccacttca
     1681 agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg
     1741 ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg
     1801 cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct
     1861 acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga
     1921 gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc
     1981 ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg
     2041 agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg
     2101 cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgttc tttcctgcgt
     2161 tatcccctga ttctgtggat aac
//
