LOCUS       sequence-242750-        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"
     protein_bind    3..27
                     /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             83..100
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     promoter        214..232
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    233..257
                     /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             1980..2024
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     primer_bind     complement(2062..2080)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2076..2123
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(2353..3168)
                     /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(2353..3168)
                     /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     3076..3095
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        complement(3169..3260)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(3276..4025)
                     /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(3286..3303)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     complement(3526..3545)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(4221..5303)
                     /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     5265..5284
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5304..5381)
                     /gene="lacI"
                     /label=lacI promoter
     promoter        join(5427..5443,1..2)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 ggggaattgt gagcggataa caattcccct gtagaaataa ttttgtttaa ctttaataag
       61 gagatatacc atgggcagca gccatcacca tcatcaccac agccaggatc cgaattcgag
      121 ctcggcgcgc ctgcaggtcg acaagcttgc ggccgcataa tgcttaagtc gaacagaaag
      181 taatcgtatt gtacacggcc gcataatcga aattaatacg actcactata ggggaattgt
      241 gagcggataa caattcccca tcttagtata ttagttaagt ataagaagga gatatacata
      301 tgttcgtgtc cgatttccgc aaagagttct acgaggtggt ccagagccag agggtccttc
      361 tcttcgtggc ctcggacgtg gatgctctgt gtgcgtgcaa gatccttcag gccttgttcc
      421 agtgtgacca cgtgcaatat acgctggttc cagtttctgg gtggcaagaa cttgaaactg
      481 catttcttga gcataaagaa cagtttcatt attttattct cataaactgt ggagctaatg
      541 tagacctatt ggatattctt caacctgatg aagacactat attctttgtg tgtgacaccc
      601 ataggccagt caatgtcgtc aatgtataca acgataccca gatcaaatta ctcattaaac
      661 aagatgatga ccttgaagtt cccgcctatg aagacatctt cagggatgaa gaggaggatg
      721 aagagcattc aggaaatgac agtgatgggt cagagcctgt ggagcaaacc atgcggagga
      781 ggcagcggcg agagtgggag gcccggagaa gagacatcct ctttgactac gagcagtatg
      841 aatatcatgg gacatcgtca gccatggtga tgtttgagct ggcttggatg ctgtccaagg
      901 acctgaatga catgctgtgg tgggccatcg ttggactaac agaccagtgg gtgcaagaca
      961 agatcactca aatgaaatac gtgactgatg ttggtgtcct gcagcgccac gtttcccgcc
     1021 acaaccaccg gaacgaggat gaggagaaca cactctccgt ggactgcaca cggatctcct
     1081 ttgagtatga cctccgcctg gtgctctacc agcactggtc cctccatgac agcctgtgca
     1141 acaccagcta taccgcagcc aggttcaagc tgtggtctgt gcatggacag aagcggctcc
     1201 aggagttcct tgcagacatg ggtcttcccc tgaagcaggt gaagcagaag ttccaggcca
     1261 tggacatctc cttgaaggag aatttgcggg aaatgattga agagtctgca aataaatttg
     1321 ggatgaagga catgcgcgtg cagactttca gcattcattt tgggttcaag cacaagtttc
     1381 tggccagcga cgtggtcttt gccaccatgt ctttgatgga gagccccgag aaggatggct
     1441 cagggacaga tcacttcatc caggctctgg acagcctctc caggagtaac ctggacaagc
     1501 tgtaccatgg cctggaactc gccaagaagc agctgcgagc cacccagcag accattgcca
     1561 gctgcctttg caccaacctc gtcatctccc aggggccttt cctgtactgc tctctcatgg
     1621 agggcactcc agatgtcatg ctgttctcta ggccggcatc cctaagcctg ctcagcaaac
     1681 acctgctcaa gtcctttgtg tgttcgacaa agaaccggcg ctgcaaactg ctgcccctgg
     1741 tgatggctgc ccccctgagc atggagcatg gcacagtgac cgtggtgggc atccccccag
     1801 agaccgacag ctcggacagg aagaactttt ttgggagggc gtttgagaag gcagcggaaa
     1861 gcaccagctc ccggatgctg cacaaccatt ttgacctctc agtaattgag ctgaaagctg
     1921 aggatcggag caagtttctg gacgcactta tttccctcct gtcctaactc gagtctggta
     1981 aagaaaccgc tgctgcgaaa tttgaacgcc agcacatgga ctcgtctact agcgcagctt
     2041 aattaaccta ggctgctgcc accgctgagc aataactagc ataacccctt ggggcctcta
     2101 aacgggtctt gaggggtttt ttgctgaaac ctcaggcatt tgagaagcac acggtcacac
     2161 tgcttccggt agtcaataaa ccggtaaacc agcaatagac ataagcggct atttaacgac
     2221 cctgccctga accgacgaca agctgacgac cgggtctccg caagtggcac ttttcgggga
     2281 aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc
     2341 atgaattaat tcttagaaaa actcatcgag catcaaatga aactgcaatt tattcatatc
     2401 aggattatca ataccatatt tttgaaaaag ccgtttctgt aatgaaggag aaaactcacc
     2461 gaggcagttc cataggatgg caagatcctg gtatcggtct gcgattccga ctcgtccaac
     2521 atcaatacaa cctattaatt tcccctcgtc aaaaataagg ttatcaagtg agaaatcacc
     2581 atgagtgacg actgaatccg gtgagaatgg caaaagttta tgcatttctt tccagacttg
     2641 ttcaacaggc cagccattac gctcgtcatc aaaatcactc gcatcaacca aaccgttatt
     2701 cattcgtgat tgcgcctgag cgagacgaaa tacgcggtcg ctgttaaaag gacaattaca
     2761 aacaggaatc gaatgcaacc ggcgcaggaa cactgccagc gcatcaacaa tattttcacc
     2821 tgaatcagga tattcttcta atacctggaa tgctgttttc ccggggatcg cagtggtgag
     2881 taaccatgca tcatcaggag tacggataaa atgcttgatg gtcggaagag gcataaattc
     2941 cgtcagccag tttagtctga ccatctcatc tgtaacatca ttggcaacgc tacctttgcc
     3001 atgtttcaga aacaactctg gcgcatcggg cttcccatac aatcgataga ttgtcgcacc
     3061 tgattgcccg acattatcgc gagcccattt atacccatat aaatcagcat ccatgttgga
     3121 atttaatcgc ggcctagagc aagacgtttc ccgttgaata tggctcatac tcttcctttt
     3181 tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg
     3241 tatttagaaa aataaacaaa taggcatgca gcgctcttcc gcttcctcgc tcactgactc
     3301 gctacgctcg gtcgttcgac tgcggcgagc ggtgtcagct cactcaaaag cggtaatacg
     3361 gttatccaca gaatcagggg ataaagccgg aaagaacatg tgagcaaaaa gcaaagcacc
     3421 ggaagaagcc aacgccgcag gcgtttttcc ataggctccg cccccctgac gagcatcaca
     3481 aaaatcgacg ctcaagccag aggtggcgaa acccgacagg actataaaga taccaggcgt
     3541 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
     3601 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgttggtatc
     3661 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc
     3721 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact
     3781 tatcgccact ggcagcagcc attggtaact gatttagagg actttgtctt gaagttatgc
     3841 acctgttaag gctaaactga aagaacagat tttggtgagt gcggtcctcc aacccactta
     3901 ccttggttca aagagttggt agctcagcga accttgagaa aaccaccgtt ggtagcggtg
     3961 gtttttcttt atttatgaga tgatgaatca atcggtctat caagtcaacg aacagctatt
     4021 ccgttactct agatttcagt gcaatttatc tcttcaaatg tagcacctga agtcagcccc
     4081 atacgatata agttgtaatt ctcatgttag tcatgccccg cgcccaccgg aaggagctga
     4141 ctgggttgaa ggctctcaag ggcatcggtc gagatcccgg tgcctaatga gtgagctaac
     4201 ttacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc
     4261 tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgccagggtg
     4321 gtttttcttt tcaccagtga gacgggcaac agctgattgc ccttcaccgc ctggccctga
     4381 gagagttgca gcaagcggtc cacgctggtt tgccccagca ggcgaaaatc ctgtttgatg
     4441 gtggttaacg gcgggatata acatgagctg tcttcggtat cgtcgtatcc cactaccgag
     4501 atgtccgcac caacgcgcag cccggactcg gtaatggcgc gcattgcgcc cagcgccatc
     4561 tgatcgttgg caaccagcat cgcagtggga acgatgccct cattcagcat ttgcatggtt
     4621 tgttgaaaac cggacatggc actccagtcg ccttcccgtt ccgctatcgg ctgaatttga
     4681 ttgcgagtga gatatttatg ccagccagcc agacgcagac gcgccgagac agaacttaat
     4741 gggcccgcta acagcgcgat ttgctggtga cccaatgcga ccagatgctc cacgcccagt
     4801 cgcgtaccgt cttcatggga gaaaataata ctgttgatgg gtgtctggtc agagacatca
     4861 agaaataacg ccggaacatt agtgcaggca gcttccacag caatggcatc ctggtcatcc
     4921 agcggatagt taatgatcag cccactgacg cgttgcgcga gaagattgtg caccgccgct
     4981 ttacaggctt cgacgccgct tcgttctacc atcgacacca ccacgctggc acccagttga
     5041 tcggcgcgag atttaatcgc cgcgacaatt tgcgacggcg cgtgcagggc cagactggag
     5101 gtggcaacgc caatcagcaa cgactgtttg cccgccagtt gttgtgccac gcggttggga
     5161 atgtaattca gctccgccat cgccgcttcc actttttccc gcgttttcgc agaaacgtgg
     5221 ctggcctggt tcaccacgcg ggaaacggtc tgataagaga caccggcata ctctgcgaca
     5281 tcgtataacg ttactggttt cacattcacc accctgaatt gactctcttc cgggcgctat
     5341 catgccatac cgcgaaaggt tttgcgccat tcgatggtgt ccgggatctc gacgctctcc
     5401 cttatgcgac tcctgcatta ggaaattaat acgactcact ata
//
