LOCUS       sequence-389079-        7854 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Heatshock expressed Cas9.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7854)
  AUTHORS   Stevenson ZC, Moerdyk-Schauwecker MJ, Banse SA, Patel DS, Lu H, 
            Phillips PC
  TITLE     High-throughput library transgenesis in Caenorhabditis elegans via 
            Transgenic Arrays Resulting in Diversity of Integrated Sequences 
            (TARDIS).
  JOURNAL   Elife. 2023 Jul 4;12:RP84831. doi: 10.7554/eLife.84831.
  PUBMED    37401921
REFERENCE   2  (bases 1 to 7854)
  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: "Elife."; 
            date: "2023-07-4"; volume: "12:RP84831. doi"; pages: " 
            10.7554/eLife.84831"
COMMENT     Sequence Label: pZCS36 (Phsp16.41::Cas9(dpiRNA)::tbb-2 '3UTR)
FEATURES             Location/Qualifiers
     source          1..7854
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    106..127
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(142..147,148..165,166..172)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    180..196
                     /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)."
     primer_bind     185..207
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     204..220
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     204..220
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             5069..5089
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     terminator      5111..5434
                     /gene="tbb-2"
                     /label=tbb-2 terminator
                     /note="transcription terminator for the C. elegans tubulin 
                     beta-2 gene"
     primer_bind     complement(5458..5475)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5458..5474)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5467..5489)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(5683..5702)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5802..5824
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5862..5880)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5948..6052
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6053..6121,6122..6913)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTSANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(6271..6290)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7084..7672
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7573..7592
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7826..7843
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca
       61 cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaatg tgagttagct
      121 cactcattag gcaccccagg ctttacactt tatgcttccg gctcgtatgt tgtgtggaat
      181 tgtgagcgga taacaatttc acacaggaaa cagctatgac catgattacg ccaagcttgc
      241 atgcctgcag gtcgactcta gaggatcccc tacaggtcga ctctagagga tcaccaaaaa
      301 cggaacgttg agctggacgg aaatagtggt aaagtgacat gattatagtt tgaagatttc
      361 taatttcaca attagagcaa atgttgttcg gtatttattt tcaacggtat ttatactatt
      421 ttccaccttt ttctagaaca ttcgagctgc ttgttgcaaa aggagggcga ctcacattcg
      481 gtacatggaa aagtagtgta cacaataaag agacccagat acattttccg tctgcgtctc
      541 tttgcaccca ccgggagtat tttcaaacga atgcatctag gaccttctag aacattctgt
      601 aaggctgcag aatgcgggta tataaggaaa gcgggctcag aggaagccaa cacgctttgt
      661 tctagtgcat ctaaaaaact tcgaaaatcc tcatcggatc ctatcgattc gcggccgctg
      721 tacacccggg tgccctatag tgagtcgtat tggtaccggt agaaaaaata tggacaaaaa
      781 atacagcatc ggactcgaca tcggtaccaa ctccgtcggt tgggccgtca tcaccgacga
      841 gtacaaggtg cgttggctaa tttcctgaat atcttcaatt tactacattt caggtcccat
      901 ccaagaagtt caaggtcctc ggtaacacag accgtcactc aatcaagaag aacctcatcg
      961 gagccctcct cttcgactcc ggagagaccg ccgaggccac ccgtctcaag cgtaccgccc
     1021 gtcgtcgtta cacccgtcgt aagaaccgta tctgctacct ccaggagatc ttctccaacg
     1081 agatggccaa ggtcgacgac tcattcttcc accgcctcga ggagtccttc ctcgtcgagg
     1141 aggacaagaa gcacgagcgt caccctatct tcggaaacat cgtcgacgag gtcgcctacc
     1201 acgagaagta cccaaccatc taccacctcc gtaagaagct cgtcgactcc accgacaagg
     1261 ccgacctccg tctcatctac ctcgccctcg cccacatgat caagttccgt ggacacttcc
     1321 tcatcgaggg agacctcaac ccagacaact ccgacgtcga caagctcttc atccagctcg
     1381 tccagaccta caaccagctc ttcgaggaga accctatcaa cgcctccgga gtcgacgcca
     1441 aggccatcct ctccgcccgt ctctccaagt cccgtcgtct cgagaacctc atcgcccagc
     1501 tcccaggaga gaagaagaac ggactcttcg gaaacctcat cgccctctcc ctcggactca
     1561 cccctaactt caagtccaac ttcgacctcg ccgaggacgc caagctccag ctctccaaag
     1621 acacctacga cgacgacctc gacaacctcc tcgcccagat cggagaccag tacgccgacc
     1681 tcttcctcgc cgccaaggtc agaagaaaac atgtattttt gctcactgta aacattttca
     1741 gaacctctcc gacgccatcc tcctctccga catcctccgt gtcaacaccg agataaccaa
     1801 ggccccactc tccgcctcca tgatcaagcg ttacgacgag caccaccagg acctcaccct
     1861 cctcaaggcc ctcgtccgtc agcagctccc agagaagtac aaggagatct tcttcgacca
     1921 gtccaagaac ggatacgccg gatacatcga cggaggagcc tcccaggagg agttctacaa
     1981 gttcatcaag ccaatcctcg aaaagatgga cggaacagag gagctcctcg tcaagctcaa
     2041 ccgtgaggac ctcctccgta agcagcgtac cttcgacaac ggatccatcc cacaccagat
     2101 ccacctcgga gagctccacg ccatcctccg tcgtcaggag gacttctacc cattcctgaa
     2161 agacaaccgc gaaaagatcg aaaagatact caccttccgt atcccatact acgtcggacc
     2221 actcgcccgt ggaaactccc gtttcgcctg gatgacccgt aagtccgagg agaccatcac
     2281 cccatggaac ttcgaggagg tcgtcgacaa gggagcctcc gcccagtcct tcatcgagcg
     2341 tatgaccaac ttcgacaaga acctccctaa cgagaaggtc ctcccaaagc actccctcct
     2401 ctacgagtac ttcaccgtct acaacgagct caccaaggtc aagtacgtca ccgagggaat
     2461 gcgtaagcca gccttcctct ccggagaaca gaagaaggcc atcgtcgacc tcctcttcaa
     2521 ggtaagttta aacatgattt tactaactaa ctaatctgat ttaaattttc agaccaaccg
     2581 taaggtgaca gtcaagcaac tcaaggagga ctacttcaag aagatcgagt gcttcgactc
     2641 cgtcgagatc tccggagtcg aggaccgttt caacgcctcc ctcggaacct accacgacct
     2701 cctcaagatc atcaaggaca aggacttcct cgacaacgag gagaacgagg acatcctcga
     2761 ggacatcgtc ctcaccctca ccctcttcga ggaccgtgag atgatcgagg agcgtctcaa
     2821 gacctacgcc cacctcttcg acgacaaggt catgaagcag ctcaagcgtc gtcgttacac
     2881 cggatgggga cgtctctccc gtaagctcat caacggaatc cgtgacaagc agtccggaaa
     2941 gaccatcctc gacttcctca agtccgacgg attcgccaac cgtaacttca tgcagctcat
     3001 ccacgacgac tccctcacct tcaaggagga catccagaag gcccaggtct ccggacaggg
     3061 agactccctc cacgagcaca tcgccaacct cgccggatcc ccagccatca agaagggaat
     3121 cctccagacc gtcaaggtcg tcgacgagct cgtcaaggtc atgggacgtc acaagccaga
     3181 gaacatcgtc atcgagatgg cccgcgagaa ccagaccacc cagaagggac agaagaactc
     3241 ccgtgagcgt atgaagcgta tcgaggaggg aatcaaggag ctcggatccc agatcctcaa
     3301 ggagcaccca gtcgagaaca cccagctcca gaacgagaag ctctacctct actacctcca
     3361 gaacggacgt gacatgtacg tcgaccagga gctggacatc aaccgtctct ccgactacga
     3421 cgtcgaccac atcgtcccac agtccttcct caaggacgac tccatagaca acaaggtgct
     3481 cacccgttcc gacaagaacc gtggaaagtc cgacaacgtc ccatccgagg aggtcgtcaa
     3541 gaagatgaag aactactggc gtcagctcct caacgccaag ctcatcaccc agcgtaagtt
     3601 cgacaacctg accaaggccg agcgtggagg actctccgag ctcgacaagg ccggattcat
     3661 caagcgtcag ctcgtcgaga cccgtcagat caccaagcac gtcgcccaga tcctcgactc
     3721 ccgtatgaac accaagtacg acgagaacga caagctcatc cgtgaggtca aggtcatcac
     3781 cctcaagtcc aagctagtct ccgacttccg taaagacttc cagttctaca aggtccgtga
     3841 gatcaacaac taccaccacg cccacgacgc ctacctcaac gccgtcgtcg gaaccgccct
     3901 catcaagaag tacccaaagc tcgagtccga gttcgtctac ggagactaca aggtctacga
     3961 cgtccgtaag gtaagaatga agcaaacaaa ttatgttaat tccaattttt agatgatcgc
     4021 caagtccgaa caggagatag gaaaggccac cgccaagtac ttcttctact ccaacatcat
     4081 gaacttcttc aagaccgaga taaccctcgc caacggagag atccgtaagc gtccactcat
     4141 cgagaccaac ggagagaccg gagagatcgt ctgggacaag ggacgtgact tcgccaccgt
     4201 ccgtaaggtc ctctccatgc cacaggtcaa catcgtcaag aagaccgagg tccagaccgg
     4261 aggattctcc aaggagtcca tcctcccaaa gcgtaactcc gacaagctca tcgcccgtaa
     4321 aaaggactgg gaccctaaga agtacggagg attcgactcc ccaaccgtcg cctactccgt
     4381 cctcgtcgtc gccaaggtcg agaagggaaa gtccaagaag ctcaagtccg tcaaggagct
     4441 cctcggaatc accatcatgg agcgttcctc cttcgagaag aacccaatcg acttcctcga
     4501 ggccaaggga tacaaggagg taaagaaaga cctcatcatc aagctcccaa agtactccct
     4561 cttcgagctc gagaacggac gtaagcgtat gctcgcctcc gccggagagc tccagaaggg
     4621 aaacgagctc gccctcccat ccaagtacgt caacttcctc tacctcgcct cccactacga
     4681 gaagctcaaa ggttccccag aagacaacga gcagaagcag ctcttcgtcg agcagcacaa
     4741 gcactacctc gacgagatca tcgagcagat ctccgagttc tccaagcgtg tcatcctcgc
     4801 cgacgccaac ctcgacaagg tcctctccgc ctacaacaag caccgtgaca agccaatccg
     4861 tgagcaggcc gagaacatca tccacctctt caccctcacc aacctcggag ccccagccgc
     4921 cttcaagtac ttcgacacca ccatcgaccg taagcgttac acctccacca aggaggtcct
     4981 cgacgccacc ctcatccacc agtccatcac cggactctac gagacccgta tcgacctctc
     5041 ccagctcgga ggagactccc gtgccgaccc aaagaagaag cgtaaggtct aattgtacaa
     5101 agtgggatcc atgcaaaatc ctttcaagca ttcccttctt ctctatcact cttctttctt
     5161 tttgtcaaaa aattctctcg ctaatttatt tgctttttta atgttattat tttatgactt
     5221 tttatagtca ctgaaaagtt tgcatctgag tgaagtgaat gctatcaaaa tgtgattctg
     5281 tctgatgtac tttcacaatc tctcttcaat tccattttga agtgctttaa acccgaaagg
     5341 ttgagaaaaa tgcgagcgct caaatatttg tattgtgttc gttgagtgac ccaacaaaaa
     5401 gaggaaactt tattgtgccg ccaagaaaaa agtctcacgg gtaccgagct cgaattcact
     5461 ggccgtcgtt ttacaacgtc gtgactggga aaaccctggc gttacccaac ttaatcgcct
     5521 tgcagcacat ccccctttcg ccagctggcg taatagcgaa gaggcccgca ccgatcgccc
     5581 ttcccaacag ttgcgcagcc tgaatggcga atggcgcctg atgcggtatt ttctccttac
     5641 gcatctgtgc ggtatttcac accgcatatg gtgcactctc agtacaatct gctctgatgc
     5701 cgcatagtta agccagcccc gacacccgcc aacacccgct gacgcgccct gacgggcttg
     5761 tctgctcccg gcatccgctt acagacaagc tgtgaccgtc tccgggagct gcatgtgtca
     5821 gaggttttca ccgtcatcac cgaaacgcgc gagacgaaag ggcctcgtga tacgcctatt
     5881 tttataggtt aatgtcatga taataatggt ttcttagacg tcaggtggca cttttcgggg
     5941 aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct
     6001 catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagtat
     6061 tcaacatttc cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc
     6121 tcacccagaa acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg
     6181 ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg
     6241 ttttccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtattga
     6301 cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta
     6361 ctcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc
     6421 tgccataacc atgagtgata acacttcggc caacttactt ctgacaacga tcggaggacc
     6481 gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg
     6541 ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc
     6601 aatggcaaca acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca
     6661 acaattaata gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct
     6721 tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat
     6781 cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg
     6841 gagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat
     6901 taagcattgg taactgtcag accaagttta ctcatatata ctttagattg atttaaaact
     6961 tcatttttaa tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat
     7021 cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc
     7081 ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct
     7141 accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg
     7201 cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca
     7261 cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc
     7321 tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga
     7381 taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac
     7441 gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga
     7501 agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag
     7561 ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg
     7621 acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag
     7681 caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc
     7741 tgcgttatcc cctgattctg tggataaccg tattaccgcc tttgagtgag ctgataccgc
     7801 tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg aaga
//
