LOCUS       sequence-71696-e        2755 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial SP crRNA expression: targets SP to protospacer B 
            (ACTTTAAAAGTATTCGCCAT), p15A/chloramphenicol.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 30 to 64)
  AUTHORS   Esvelt KM, Mali P, Braff JL, Moosburner M, Yaung SJ, Church GM
  TITLE     Orthogonal Cas9 proteins for RNA-guided gene regulation and editing.
  JOURNAL   Nat Methods. 2013 Sep 29. doi: 10.1038/nmeth.2681.
  PUBMED    24076762
REFERENCE   2  (bases 1 to 2755)
  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: "Nat 
            Methods. 2013 Sep 29. doi: 10.1038/nmeth.2681."
COMMENT     Sequence Label: PM-SP!TB
FEATURES             Location/Qualifiers
     source          1..2755
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        30..64
                     /label=﻿BBa_J23100
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 1.0
                     "
                     /citation=[1]
     repeat_region   106..141
                     /label=DR
                     /note="direct repeat for the Streptococcus pyogenes 
                     CRISPR/Cas system"
     terminator      complement(239..266)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(358..444)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     promoter        665..767
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     CDS             768..1427
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(834..853)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     terminator      complement(1437..1523)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     complement(1813..1830)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1947..2492)
                     /direction=LEFT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     complement(2027..2046)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gacaatgaaa acgttagtca tggcgcgcct tgacggctag ctcagtccta ggtacagtgc
       61 tagcttaatg ctcgcacata gcagaacttt aaaagtattc gccatgtttt agagctatgc
      121 tgttttgaat ggtcccaaaa cccgccgcta tggtcccacg tagagcatac ggaaaaaaaa
      181 gtcaaaagcc tccgaccgga ggtcggcctt acttgctagc agagtttgta gaaacgcaaa
      241 aaggccatcc gtcaggatgg ccttctgctt aatttgatgc ctggcagttt atggcgggcg
      301 tcctgcccgc caccctccgg gccgttgctt cgcaacgttc aaatccgctc ccggcggatt
      361 tgtcctactc aggagagcgt tcaccgacaa acaacagata aaacgaaagg cccagtcttt
      421 cgactgagcc tttcgtttta tttgatgcct ggcagttccc tactctcgca tggggagacc
      481 ccacactacc atcggcgcta cggcgtttca cttctgagtt cggcatgggg tcaggtggga
      541 ccaccgcgct actgccgcca ggcaaattct gttttatcag accgcttctg cgttctgatt
      601 taatctgtat caggctgaaa atcttctctc atccgccaaa acagccccgt agaaaaaggg
      661 acgttgatcg gcacgtaaga ggttccaact ttcaccataa tgaaataaga tcactaccgg
      721 gcgtattttt tgagttatcg agattttcag gagctaagga agctaaaatg gagaaaaaaa
      781 tcactggata taccaccgtt gatatatccc aatggcatcg taaagaacat tttgaggcat
      841 ttcagtcagt tgctcaatgt acctataacc agaccgttca gctggatatt acggcctttt
      901 taaagaccgt aaagaaaaat aagcacaagt tttatccggc ctttattcac attcttgccc
      961 gcctgatgaa tgctcatccg gaattccgta tggcaatgaa agacggtgag ctggtgatat
     1021 gggatagtgt tcacccttgt tacaccgttt tccatgagca aactgaaacg ttttcatcgc
     1081 tctggagtga ataccacgac gatttccggc agtttctaca catatattcg caagatgtgg
     1141 cgtgttacgg tgaaaacctg gcctatttcc ctaaagggtt tattgagaat atgtttttcg
     1201 tctcagccaa tccctgggtg agtttcacca gttttgattt aaacgtggcc aatatggaca
     1261 acttcttcgc ccccgttttc accatgggca aatattatac gcaaggcgac aaggtgctga
     1321 tgccgctggc gattcaggtt catcatgccg tctgtgatgg cttccatgtc ggcagaatgc
     1381 ttaatgaatt acaacagtac tgcgatgagt ggcagggcgg ggcgtaattc cggcggattt
     1441 gtcctactca ggagagcgtt caccgacaaa caacagataa aacgaaaggc ccagtctttc
     1501 gactgagcct ttcgttttat ttgatgcctg gcagttccct actctcgcat ggggagaccc
     1561 cacactacca tcggcgctac ggcgtttcac ttctgagttc ggcatggggt caggtgggac
     1621 caccgcgcta ctgccgccag gcaaattctg ttttatcaga ccgcttctgc gttctgattt
     1681 aatctgtatc aggctgaaaa tcttctctca tccgccaaaa cagccctcgg gacggatcgc
     1741 ttcatgtggc aggagaaaaa aggctgcacc ggtgcgtcag cagaatatgt gatacaggat
     1801 atattccgct tcctcgctca ctgactcgct acgctcggtc gttcgactgc ggcgagcgga
     1861 aatggcttac gaacggggcg gagatttcct ggaagatgcc aggaagatac ttaacaggga
     1921 agtgagaggg ccgcggcaaa gccgtttttc cataggctcc gcccccctga caagcatcac
     1981 gaaatctgac gctcaaatca gtggtggcga aacccgacag gactataaag ataccaggcg
     2041 tttccccctg gcggctccct cgtgcgctct cctgttcctg cctttcggtt taccggtgtc
     2101 attccgctgt tatggccgcg tttgtctcat tccacgcctg acactcagtt ccgggtaggc
     2161 agttcgctcc aagctggact gtatgcacga accccccgtt cagtccgacc gctgcgcctt
     2221 atccggtaac tatcgtcttg agtccaaccc ggaaagacat gcaaaagcac cactggcagc
     2281 agccactggt aattgattta gaggagttag tcttgaagtc atgcgccggt taaggctaaa
     2341 ctgaaaggac aagttttggt gactgcgctc ctccaagcca gttacctcgg ttcaaagagt
     2401 tggtagctca gagaaccttc gaaaaaccgc cctgcaaggc ggttttttcg ttttcagagc
     2461 aagagattac gcgcagacca aaacgatctc aagaagatca tcttattaag gggtctgacg
     2521 ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct
     2581 tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt
     2641 aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca gcggaaccgc
     2701 tctagatttc agtgcaattt atctcttcaa atgtagcacc tgaagtcagc ccggc
//
