LOCUS       sequence-257710-        2470 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Engineered sgRNA-LEB1 generator including Anderson promoter J23100 
            for golden gate assembly.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 343 to 377)
  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 Dec 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: pLY130
FEATURES             Location/Qualifiers
     source          1..2470
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     69..86
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(110..167)
                     /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."
     misc_feature    168..188
                     /label=BioBrick suffix
                     /note="universal suffix for all parts"
     protein_bind    232..250
                     /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    287..305
                     /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)"
     promoter        complement(343..377)
                     /label=﻿BBa_J23100
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 1.0
                     "
                     /citation=[1]
     misc_feature    455..476
                     /label=BioBrick prefix
                     /note="BioBrick prefix for parts that do not start with 
                     ""ATG"""
     terminator      479..522
                     /label=bacterial terminator
                     /note="putative bacterial transcription terminator"
     primer_bind     complement(526..544)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(738..757)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             749..1564
                     /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             749..1564
                     /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     complement(822..841)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      1797..2385
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2286..2305
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga
       61 ccgagcgcag cgagtcagtg agcgaggaag cctgcataac gcgaagtaat cttttcggtt
      121 ttaaagaaaa agggcagggt ggtgacacct tgcccttttt tgccggactg cagcggccgc
      181 tactagtaga agacaagcta cttcaaaaaa agcaccgact cggtgccact tgggcccttc
      241 ttcagggccc aagttgataa cggactagcc ttatttgaac ttgctagggc ccttcttcag
      301 ggccctagct ctcaaacatc ttccagacct cagactctag tagctagcac tgtacctagg
      361 actgagctag ccgtcaactc tagtaccttt ggtcgaaaaa aaaagcccgc actgtcaggt
      421 gcgggctttt ttctgtgttt ccaattttgt cttcctctag aagcggccgc gaattccaga
      481 aatcatcctt agcgaaagct aaggattttt tttatctgaa attctgcctc gtgatacgcc
      541 tatttttata ggttaatgtc atgataataa tggtttctta gacgtcaagt ggcactcgag
      601 ctgatccttc aactcagcaa aagttcgatt tattcaacaa agccacgttg tgtctcaaaa
      661 tctctgatgt tacattgcac aagataaaaa tatatcatca tgaacaataa aactgtctgc
      721 ttacataaac agtaatacaa ggggtgttat gagccatatt caacgggaaa cgtcttgctc
      781 caggccgcga ttaaattcca acatggatgc tgatttatat gggtataaat gggctcgcga
      841 taatgtcggg caatcaggtg cgacaatcta tcgattgtat gggaagcccg atgcgccaga
      901 gttgtttctg aaacatggca aaggtagcgt tgccaatgat gttacagatg agatggtcag
      961 actaaactgg ctgacggaat ttatgcctct tccgaccatc aagcatttta tccgtactcc
     1021 tgatgatgca tggttactca ccactgcgat ccccgggaaa acagcattcc aggtattaga
     1081 agaatatcct gattcaggtg aaaatattgt tgatgcgctg gcagtgttcc tgcgccggtt
     1141 gcattcgatt cctgtttgta attgtccttt taacagcgat cgcgtatttc gtctcgctca
     1201 ggcgcaatca cgaatgaata acggtttggt tgatgcgagt gattttgatg acgagcgtaa
     1261 tggctggcct gttgaacaag tctggaaaga aatgcataag cttttgccat tctcaccgga
     1321 ttcagtcgtc actcatggtg atttctcact tgataacctt atttttgacg aggggaaatt
     1381 aataggttgt attgatgttg gacgagtcgg aatcgcagac cgataccagg atcttgccat
     1441 cctatggaac tgcctcggtg agttttctcc ttcattacag aaacggcttt ttcaaaaata
     1501 tggtattgat aatcctgata tgaataaatt gcagtttcat ttgatgctcg atgagttttt
     1561 ctaatcagaa ttggttaatt ggttgtaaca ctggcagagc attacgctga cttgacggga
     1621 ctcgagctgt cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt
     1681 taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa
     1741 cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga
     1801 gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg
     1861 gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc
     1921 agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag
     1981 aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc
     2041 agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg
     2101 cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac
     2161 accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga
     2221 aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt
     2281 ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag
     2341 cgtcgatttt tgtgatgctc gtcagggggg cggagcctgt ggaaaaacgc cagcaacgcg
     2401 gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta
     2461 tcccctgatt
//
