LOCUS       sequence-257712-        2470 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Engineered sgRNA-LEB3 generator including Anderson promoter J23100 
            for golden gate assembly.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 196 to 230)
  AUTHORS   Liu Y, Wan X, Wang B
  TITLE     Engineered CRISPRa enables programmable eukaryote-like gene 
            activation in bacteria.
  JOURNAL   Nat Commun. 2019 Aug 26;10(1):3693. doi: 10.1038/s41467-019-11479-0.
  PUBMED    31451697
REFERENCE   2  (bases 1 to 2470)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Commun."; date: "2019-08-26"; volume: "10(1):3693. doi"; pages: " 
            10.1038/s41467-019-11479-0"
COMMENT     Sequence Label: pLY132
FEATURES             Location/Qualifiers
     source          1..2470
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     29..47
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     terminator      complement(51..94)
                     /label=bacterial terminator
                     /note="putative bacterial transcription terminator"
     misc_feature    97..118
                     /label=BioBrick prefix
                     /note="BioBrick prefix for parts that do not start with 
                     ""ATG"""
     promoter        196..230
                     /label=﻿BBa_J23100
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 1.0
                     "
                     /citation=[1]
     protein_bind    268..286
                     /label=boxB
                     /bound_moiety="lambda-N peptide"
                     /note="lambda N peptide binding site consisting of an RNA 
                     hairpin within the N-utliization (nut) site of the lambda 
                     bacteriophage transcript (Tan and Frankel, 1995)"
     protein_bind    323..341
                     /label=boxB
                     /bound_moiety="lambda-N peptide"
                     /note="lambda N peptide binding site consisting of an RNA 
                     hairpin within the N-utliization (nut) site of the lambda 
                     bacteriophage transcript (Tan and Frankel, 1995)"
     misc_feature    385..405
                     /label=BioBrick suffix
                     /note="universal suffix for all parts"
     terminator      406..463
                     /label=his operon terminator
                     /note="This putative transcriptin terminator from the E. 
                     coli his operon has a 2-bp deletion introduced during 
                     synthesis. Its efficiency has not been determined."
     primer_bind     complement(487..504)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(658..1246)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(738..757)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(1479..2294)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(1479..2294)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     2202..2221
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2286..2305
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
ORIGIN
        1 cattattatc atgacattaa cctataaaaa taggcgtatc acgaggcaga atttcagata
       61 aaaaaaatcc ttagctttcg ctaaggatga tttctggaat tcgcggccgc ttctagagga
      121 agacaaccta ggaaacacag aaaaaagccc gcacctgaca gtgcgggctt tttttttcga
      181 ccaaaggtac tagagttgac ggctagctca gtcctaggta cagtgctagc tactagagca
      241 tagttcgttt cccatgtttg agagctaggg ccctgaagaa gggccctagc aagttcaaat
      301 aaggctagtc cgttatcaac ttgggccctg aagaagggcc caagtggcac cgagtcggtg
      361 ctttttttaa agacgtttgt cttctactag tagcggccgc tgcagtccgg caaaaaaggg
      421 caaggtgtca ccaccctgcc ctttttcttt aaaaccgaaa agattacttc gcgttatgca
      481 ggcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc
      541 tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat
      601 gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt
      661 ccacaggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg
      721 aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc
      781 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt
      841 ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa
      901 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta
      961 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa
     1021 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa
     1081 ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt
     1141 cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt
     1201 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat
     1261 cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat
     1321 gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc
     1381 aatctaaagt atatatgagt aaacttggtc tgacagctcg agtcccgtca agtcagcgta
     1441 atgctctgcc agtgttacaa ccaattaacc aattctgatt agaaaaactc atcgagcatc
     1501 aaatgaaact gcaatttatt catatcagga ttatcaatac catatttttg aaaaagccgt
     1561 ttctgtaatg aaggagaaaa ctcaccgagg cagttccata ggatggcaag atcctggtat
     1621 cggtctgcga ttccgactcg tccaacatca atacaaccta ttaatttccc ctcgtcaaaa
     1681 ataaggttat caagtgagaa atcaccatga gtgacgactg aatccggtga gaatggcaaa
     1741 agcttatgca tttctttcca gacttgttca acaggccagc cattacgctc gtcatcaaaa
     1801 tcactcgcat caaccaaacc gttattcatt cgtgattgcg cctgagcgag acgaaatacg
     1861 cgatcgctgt taaaaggaca attacaaaca ggaatcgaat gcaaccggcg caggaacact
     1921 gccagcgcat caacaatatt ttcacctgaa tcaggatatt cttctaatac ctggaatgct
     1981 gttttcccgg ggatcgcagt ggtgagtaac catgcatcat caggagtacg gataaaatgc
     2041 ttgatggtcg gaagaggcat aaattccgtc agccagttta gtctgaccat ctcatctgta
     2101 acatcattgg caacgctacc tttgccatgt ttcagaaaca actctggcgc atcgggcttc
     2161 ccatacaatc gatagattgt cgcacctgat tgcccgacat tatcgcgagc ccatttatac
     2221 ccatataaat cagcatccat gttggaattt aatcgcggcc tggagcaaga cgtttcccgt
     2281 tgaatatggc tcataacacc ccttgtatta ctgtttatgt aagcagacag ttttattgtt
     2341 catgatgata tatttttatc ttgtgcaatg taacatcaga gattttgaga cacaacgtgg
     2401 ctttgttgaa taaatcgaac ttttgctgag ttgaaggatc agctcgagtg ccacttgacg
     2461 tctaagaaac
//
