LOCUS       sequence-245620-        5443 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses crystallisation construct of human Cdc45 in bacteria.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5443)
  AUTHORS   Simon AC, Sannino V, Costanzo V, Pellegrini L
  TITLE     Structure of human Cdc45 and implications for CMG helicase function.
  JOURNAL   Nat Commun. 2016 May 18;7:11638. doi: 10.1038/ncomms11638.
  PUBMED    27189187
REFERENCE   2  (bases 1 to 5443)
  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: "2016-05-18"; volume: "7:11638. doi"; pages: " 
            10.1038/ncomms11638"
COMMENT     Sequence Label: pRSFDuet1-Cdc45 (delta 154–164)
FEATURES             Location/Qualifiers
     source          1..5443
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        114..132
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    133..157
                     /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)."
     CDS             1880..1924
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     primer_bind     complement(1962..1980)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1976..2023
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(2253..3068)
                     /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(2253..3068)
                     /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     2976..2995
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        complement(3069..3160)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(3176..3925)
                     /direction=LEFT
                     /label=RSF ori
                     /note="Plasmids containing the RSF 1030 origin of 
                     replication can be propagated in E. coli cells that contain
                     additional plasmids with compatible origins."
     primer_bind     complement(3186..3203)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     complement(3426..3445)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(4121..5203)
                     /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     5165..5184
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5204..5281)
                     /gene="lacI"
                     /label=lacI promoter
     promoter        5327..5345
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    5346..5370
                     /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)."
     CDS             5426..5443
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
ORIGIN
        1 agccaggatc cgaattcgag ctcggcgcgc ctgcaggtcg acaagcttgc ggccgcataa
       61 tgcttaagtc gaacagaaag taatcgtatt gtacacggcc gcataatcga aattaatacg
      121 actcactata ggggaattgt gagcggataa caattcccca tcttagtata ttagttaagt
      181 ataagaagga gatatacata tgttcgtgtc cgatttccgc aaagagttct acgaggtggt
      241 ccagagccag agggtccttc tcttcgtggc ctcggacgtg gatgctctgt gtgcgtgcaa
      301 gatccttcag gccttgttcc agtgtgacca cgtgcaatat acgctggttc cagtttctgg
      361 gtggcaagaa cttgaaactg catttcttga gcataaagaa cagtttcatt attttattct
      421 cataaactgt ggagctaatg tagacctatt ggatattctt caacctgatg aagacactat
      481 attctttgtg tgtgacaccc ataggccagt caatgtcgtc aatgtataca acgataccca
      541 gatcaaatta ctcattaaac aagatgatga ccttgaagtt cccgcctatg aagacatctt
      601 cagggatgaa gaggaggatg aagagcattc aggaaatgac agtgatgggt cagagcctgt
      661 ggagcaaacc atgcggagga ggcagcggcg agagtgggag gcccggagaa gagacatcct
      721 ctttgactac gagcagtatg aatatcatgg gacatcgtca gccatggtga tgtttgagct
      781 ggcttggatg ctgtccaagg acctgaatga catgctgtgg tgggccatcg ttggactaac
      841 agaccagtgg gtgcaagaca agatcactca aatgaaatac gtgactgatg ttggtgtcct
      901 gcagcgccac gtttcccgcc acaaccaccg gaacgaggat gaggagaaca cactctccgt
      961 ggactgcaca cggatctcct ttgagtatga cctccgcctg gtgctctacc agcactggtc
     1021 cctccatgac agcctgtgca acaccagcta taccgcagcc aggttcaagc tgtggtctgt
     1081 gcatggacag aagcggctcc aggagttcct tgcagacatg ggtcttcccc tgaagcaggt
     1141 gaagcagaag ttccaggcca tggacatctc cttgaaggag aatttgcggg aaatgattga
     1201 agagtctgca aataaatttg ggatgaagga catgcgcgtg cagactttca gcattcattt
     1261 tgggttcaag cacaagtttc tggccagcga cgtggtcttt gccaccatgt ctttgatgga
     1321 gagccccgag aaggatggct cagggacaga tcacttcatc caggctctgg acagcctctc
     1381 caggagtaac ctggacaagc tgtaccatgg cctggaactc gccaagaagc agctgcgagc
     1441 cacccagcag accattgcca gctgcctttg caccaacctc gtcatctccc aggggccttt
     1501 cctgtactgc tctctcatgg agggcactcc agatgtcatg ctgttctcta ggccggcatc
     1561 cctaagcctg ctcagcaaac acctgctcaa gtcctttgtg tgttcgacaa agaaccggcg
     1621 ctgcaaactg ctgcccctgg tgatggctgc ccccctgagc atggagcatg gcacagtgac
     1681 cgtggtgggc atccccccag agaccgacag ctcggacagg aagaactttt ttgggagggc
     1741 gtttgagaag gcagcggaaa gcaccagctc ccggatgctg cacaaccatt ttgacctctc
     1801 agtaattgag ctgaaagctg aggatcggag caagtttctg gacgcactta tttccctcct
     1861 gtcctaactc gagtctggta aagaaaccgc tgctgcgaaa tttgaacgcc agcacatgga
     1921 ctcgtctact agcgcagctt aattaaccta ggctgctgcc accgctgagc aataactagc
     1981 ataacccctt ggggcctcta aacgggtctt gaggggtttt ttgctgaaac ctcaggcatt
     2041 tgagaagcac acggtcacac tgcttccggt agtcaataaa ccggtaaacc agcaatagac
     2101 ataagcggct atttaacgac cctgccctga accgacgaca agctgacgac cgggtctccg
     2161 caagtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac
     2221 attcaaatat gtatccgctc atgaattaat tcttagaaaa actcatcgag catcaaatga
     2281 aactgcaatt tattcatatc aggattatca ataccatatt tttgaaaaag ccgtttctgt
     2341 aatgaaggag aaaactcacc gaggcagttc cataggatgg caagatcctg gtatcggtct
     2401 gcgattccga ctcgtccaac atcaatacaa cctattaatt tcccctcgtc aaaaataagg
     2461 ttatcaagtg agaaatcacc atgagtgacg actgaatccg gtgagaatgg caaaagttta
     2521 tgcatttctt tccagacttg ttcaacaggc cagccattac gctcgtcatc aaaatcactc
     2581 gcatcaacca aaccgttatt cattcgtgat tgcgcctgag cgagacgaaa tacgcggtcg
     2641 ctgttaaaag gacaattaca aacaggaatc gaatgcaacc ggcgcaggaa cactgccagc
     2701 gcatcaacaa tattttcacc tgaatcagga tattcttcta atacctggaa tgctgttttc
     2761 ccggggatcg cagtggtgag taaccatgca tcatcaggag tacggataaa atgcttgatg
     2821 gtcggaagag gcataaattc cgtcagccag tttagtctga ccatctcatc tgtaacatca
     2881 ttggcaacgc tacctttgcc atgtttcaga aacaactctg gcgcatcggg cttcccatac
     2941 aatcgataga ttgtcgcacc tgattgcccg acattatcgc gagcccattt atacccatat
     3001 aaatcagcat ccatgttgga atttaatcgc ggcctagagc aagacgtttc ccgttgaata
     3061 tggctcatac tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg
     3121 agcggataca tatttgaatg tatttagaaa aataaacaaa taggcatgca gcgctcttcc
     3181 gcttcctcgc tcactgactc gctacgctcg gtcgttcgac tgcggcgagc ggtgtcagct
     3241 cactcaaaag cggtaatacg gttatccaca gaatcagggg ataaagccgg aaagaacatg
     3301 tgagcaaaaa gcaaagcacc ggaagaagcc aacgccgcag gcgtttttcc ataggctccg
     3361 cccccctgac gagcatcaca aaaatcgacg ctcaagccag aggtggcgaa acccgacagg
     3421 actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac
     3481 cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca
     3541 tagctcacgc tgttggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt
     3601 gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc
     3661 caacccggta agacacgact tatcgccact ggcagcagcc attggtaact gatttagagg
     3721 actttgtctt gaagttatgc acctgttaag gctaaactga aagaacagat tttggtgagt
     3781 gcggtcctcc aacccactta ccttggttca aagagttggt agctcagcga accttgagaa
     3841 aaccaccgtt ggtagcggtg gtttttcttt atttatgaga tgatgaatca atcggtctat
     3901 caagtcaacg aacagctatt ccgttactct agatttcagt gcaatttatc tcttcaaatg
     3961 tagcacctga agtcagcccc atacgatata agttgtaatt ctcatgttag tcatgccccg
     4021 cgcccaccgg aaggagctga ctgggttgaa ggctctcaag ggcatcggtc gagatcccgg
     4081 tgcctaatga gtgagctaac ttacattaat tgcgttgcgc tcactgcccg ctttccagtc
     4141 gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt
     4201 gcgtattggg cgccagggtg gtttttcttt tcaccagtga gacgggcaac agctgattgc
     4261 ccttcaccgc ctggccctga gagagttgca gcaagcggtc cacgctggtt tgccccagca
     4321 ggcgaaaatc ctgtttgatg gtggttaacg gcgggatata acatgagctg tcttcggtat
     4381 cgtcgtatcc cactaccgag atgtccgcac caacgcgcag cccggactcg gtaatggcgc
     4441 gcattgcgcc cagcgccatc tgatcgttgg caaccagcat cgcagtggga acgatgccct
     4501 cattcagcat ttgcatggtt tgttgaaaac cggacatggc actccagtcg ccttcccgtt
     4561 ccgctatcgg ctgaatttga ttgcgagtga gatatttatg ccagccagcc agacgcagac
     4621 gcgccgagac agaacttaat gggcccgcta acagcgcgat ttgctggtga cccaatgcga
     4681 ccagatgctc cacgcccagt cgcgtaccgt cttcatggga gaaaataata ctgttgatgg
     4741 gtgtctggtc agagacatca agaaataacg ccggaacatt agtgcaggca gcttccacag
     4801 caatggcatc ctggtcatcc agcggatagt taatgatcag cccactgacg cgttgcgcga
     4861 gaagattgtg caccgccgct ttacaggctt cgacgccgct tcgttctacc atcgacacca
     4921 ccacgctggc acccagttga tcggcgcgag atttaatcgc cgcgacaatt tgcgacggcg
     4981 cgtgcagggc cagactggag gtggcaacgc caatcagcaa cgactgtttg cccgccagtt
     5041 gttgtgccac gcggttggga atgtaattca gctccgccat cgccgcttcc actttttccc
     5101 gcgttttcgc agaaacgtgg ctggcctggt tcaccacgcg ggaaacggtc tgataagaga
     5161 caccggcata ctctgcgaca tcgtataacg ttactggttt cacattcacc accctgaatt
     5221 gactctcttc cgggcgctat catgccatac cgcgaaaggt tttgcgccat tcgatggtgt
     5281 ccgggatctc gacgctctcc cttatgcgac tcctgcatta ggaaattaat acgactcact
     5341 ataggggaat tgtgagcgga taacaattcc cctgtagaaa taattttgtt taactttaat
     5401 aaggagatat accatgggca gcagccatca ccatcatcac cac
//
