LOCUS       sequence-229421-        2183 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Encodes sgRNA targeting position 5 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_pos5_AmpR
FEATURES             Location/Qualifiers
     source          1..2183
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        68..102
                     /label=J23119 promoter
                     /note="bacterial promoter (Registry of Standard Biological 
                     Parts BBa_J23119)"
     misc_RNA        123..198
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     CDS             complement(join(460..1251,1252..1320))
                     /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     1083..1102
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1321..1425)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      1528..2116
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2017..2036
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ctgcgttatc ccctgattct gtggataacc gtgcggcccg agagcatcga tcctagcatg
       61 gatgcatttg acagctagct cagtcctagg tataatgcta gctgttagct cagcatgccc
      121 aggttttaga gctagaaata gcaagttaaa ataaggctag tccgttatca acttgaaaaa
      181 gtggcaccga gtcggtgctt tttttccggc ttatcggtca gtttcacctg atttacgtaa
      241 aaacccgctt cggcgggttt ttgcttttgg aggggcagaa agatgaatga ctgtccacga
      301 cgctataccc aaaagaaaaa gcttgaaaga atcgatcgat cccgaatgag tagtacagcg
      361 gccgcgatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa
      421 atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga
      481 ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt
      541 gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg
      601 ggacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga
      661 gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga
      721 agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg
      781 catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc
      841 aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc
      901 gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca
      961 taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac
     1021 caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg
     1081 ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc
     1141 ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg
     1201 tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac
     1261 aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat
     1321 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata
     1381 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa
     1441 agtgccacct gtcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga
     1501 ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg
     1561 cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc
     1621 aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct
     1681 agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc
     1741 tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt
     1801 ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg
     1861 cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct
     1921 atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag
     1981 ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag
     2041 tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg
     2101 gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg
     2161 gccttttgct cacatgttct ttc
//
