LOCUS       sequence-306385-        3578 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of the sgRNA targeting to the E. coli can gene.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3578)
  AUTHORS   Tan SI, Yu PJ, Ng IS
  TITLE     CRISPRi-mediated Programming Essential Gene can as a Direct 
  JOURNAL   Biotechnol Bioeng. 2020 May 27. doi: 10.1002/bit.27443.
  PUBMED    32458463
REFERENCE   2  (bases 1 to 3578)
  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: "Biotechnol 
            Bioeng. 2020 May 27. doi: 10.1002/bit.27443."
COMMENT     Sequence Label: pSUI-SynP-CAS2sg
FEATURES             Location/Qualifiers
     source          1..3578
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     120..138
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     terminator      complement(142..185)
                     /label=bacterial terminator
                     /note="putative bacterial transcription terminator"
     misc_feature    188..209
                     /label=BioBrick prefix
                     /note="BioBrick prefix for parts that do not start with 
                     ""ATG"""
     protein_bind    213..229
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     misc_RNA        287..362
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     misc_feature    369..389
                     /label=BioBrick suffix
                     /note="universal suffix for all parts"
     promoter        436..513
                     /gene="lacI"
                     /label=lacI promoter
     CDS             514..1596
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=lacI
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     complement(533..552)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     primer_bind     complement(1794..1811)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1965..2553)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2045..2064)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(2877..3536)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     3451..3470
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
ORIGIN
        1 tacgcccggt agtgatctta tttcattatg gtgaaagttg gaacctctta cgtgcccgat
       61 caactcgagt gccacctgac gtctaagaaa ccattattat catgacatta acctataaaa
      121 ataggcgtat cacgaggcag aatttcagat aaaaaaaatc cttagctttc gctaaggatg
      181 atttctggaa ttcgcggccg cttctagagg aattgtgagc ggataacaat tttgacggtg
      241 tgagcggata acagtacagt ggatccatct gtggccacta cggttggttt tagagctaga
      301 aatagcaagt taaaataagg ctagtccgtt atcaacttga aaaagtggca ccgagtcggt
      361 gcttttttta ctagtagcgg ccgctgcagt gcttcctaat gcaggagtcg cataagggag
      421 agcgtcgaga tcccggacac catcgaatgg cgcaaaacct ttcgcggtat ggcatgatag
      481 cgcccggaag agagtcaatt cagggtggtg aatgtgaaac cagtaacgtt atacgatgtc
      541 gcagagtatg ccggtgtctc ttatcagacc gtttcccgcg tggtgaacca ggccagccac
      601 gtttctgcga aaacgcggga aaaagtggaa gcggcgatgg cggagctgaa ttacattccc
      661 aaccgcgtgg cacaacaact ggcgggcaaa cagtcgttgc tgattggcgt tgccacctcc
      721 agtctggccc tgcacgcgcc gtcgcaaatt gtcgcggcga ttaaatctcg cgccgatcaa
      781 ctgggtgcca gcgtggtggt gtcgatggta gaacgaagcg gcgtcgaagc ctgtaaagcg
      841 gcggtgcaca atcttctcgc gcaacgcgtc agtgggctga tcattaacta tccgctggat
      901 gaccaggatg ccattgctgt ggaagctgcc tgcactaatg ttccggcgtt atttcttgat
      961 gtctctgacc agacacccat caacagtatt attttctccc atgaagacgg tacgcgactg
     1021 ggcgtggagc atctggtcgc attgggtcac cagcaaatcg cgctgttagc gggcccatta
     1081 agttctgtct cggcgcgtct gcgtctggct ggctggcata aatatctcac tcgcaatcaa
     1141 attcagccga tagcggaacg ggaaggcgac tggagtgcca tgtccggttt tcaacaaacc
     1201 atgcaaatgc tgaatgaggg catcgttccc actgcgatgc tggttgccaa cgatcagatg
     1261 gcgctgggcg caatgcgcgc cattaccgag tccgggctgc gcgttggtgc ggatatctcg
     1321 gtagtgggat acgacgatac cgaagacagc tcatgttata tcccgccgtt aaccaccatc
     1381 aaacaggatt ttcgcctgct ggggcaaacc agcgtggacc gcttgctgca actctctcag
     1441 ggccaggcgg tgaagggcaa tcagctgttg cccgtctcac tggtgaaaag aaaaaccacc
     1501 ctggcgccca atacgcaaac cgcctctccc cgcgcgttgg ccgattcatt aatgcagctg
     1561 gcacgacagg tttcccgact ggaaagcggg cagtgagcgc aacgcaatta atgtaagtta
     1621 gctcactcat taggcaccgg gatctcgacc gatgcccttg agagccttca acccagtcag
     1681 ctccttccgg tgggcgcggg gcatgactat cgtcgccgca cttatgagat ctattaatga
     1741 atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc
     1801 actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg
     1861 gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc
     1921 cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc
     1981 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga
     2041 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc
     2101 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat
     2161 agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg
     2221 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc
     2281 aacccggtaa gacacgactt atcgccaccg gcagcagcca ctggtaacag gattagcaga
     2341 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact
     2401 agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt
     2461 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag
     2521 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg
     2581 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa
     2641 aggatcttca cctagatcct cttaaattaa aaatgaagtt ttaaatcaat ctaaagtata
     2701 tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg
     2761 atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata
     2821 cgggagggct taccatctgg ctccagtgct gcaatgatac cgcgagagtc gacaaattac
     2881 gccccgccct gccactcatc gcagtactgt tgtaattcat taagcattct gccgacatgg
     2941 aagccatcac aaacggcatg atgaacctga atcgccagcg gcatcagcac cttgtcgcct
     3001 tgcgtataat atttgcccat ggtgaaaacg ggggcgaaga agttgtccat attggccacg
     3061 tttaaatcaa aactggtgaa actcacccag ggattggctg agacgaaaaa catattctca
     3121 ataaaccctt tagggaaata ggccaggttt tcaccgtaac acgccacatc ttgcgaatat
     3181 atgtgtagaa actgccggaa atcgtcgtgg tattcactcc agagcgatga aaacgtttca
     3241 gtttgctcat ggaaaacggt gtaacaaggg tgaacactat cccatatcac cagctcaccg
     3301 tctttcattg ccatacgaaa ttccggatga gcattcatca ggcgggcaag aatgtgaata
     3361 aaggccggat aaaacttgtg cttatttttc tttacggtct ttaaaaaggc cgtaatatcc
     3421 agctgaacgg tctggttata ggtacattga gcaactgact gaaatgcctc aaaatgttct
     3481 ttacgatgcc attgggatat atcaacggtg gtatatccag tgattttttt ctccatttta
     3541 gcttccttag ctcctgaaaa tctcgataac tcaaaaaa
//
