LOCUS       sequence-320016-        5288 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses GST-tagged S.pombe Suc1 in E.coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5288)
  AUTHORS   Singh P, Halova L, Hagan IM
  TITLE     Highly Synchronous Mitotic Progression in Schizosaccharomyces pombe 
            Upon Relief of Transient Cdc2-asM17 Inhibition.
  JOURNAL   Methods Mol Biol. 2021;2329:123-142. doi: 
            10.1007/978-1-0716-1538-6_10.
  PUBMED    34085220
REFERENCE   2  (bases 1 to 5288)
  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: "Methods Mol
            Biol."; date: "2021"; volume: "2329:123-142. doi"; pages: " 
            10.1007/978-1-0716-1538-6_10"
COMMENT     Sequence Label: pGEX-4T-3-SpSuc1
FEATURES             Location/Qualifiers
     source          1..5288
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(180..185,186..201,202..208)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    216..232
                     /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             255..908
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     866..888
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             915..932
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     primer_bind     complement(1335..1357)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1502..1520)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1588..1692
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1693..1761,1762..2553)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(1911..1930)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2724..3312
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3213..3232
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3466..3483
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        3556..3633
                     /gene="lacI"
                     /label=lacI promoter
     CDS             3634..4716
                     /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(3653..3672)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(4765..4770,4771..4788,4789..4795)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4803..4819
                     /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     4827..4843
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4827..4843
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             join(4839..5288,1..78)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(4859..4876)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4859..4875)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4868..4890)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 ttatcgactg cacggtgcac caatgcttct ggcgtcaggc agccatcgga agctgtggta
       61 tggctgtgca ggtcgtaaat cactgcataa ttcgtgtcgc tcaaggcgca ctcccgttct
      121 ggataatgtt ttttgcgccg acatcataac ggttctggca aatattctga aatgagctgt
      181 tgacaattaa tcatcggctc gtataatgtg tggaattgtg agcggataac aatttcacac
      241 aggaaacagt attcatgtcc cctatactag gttattggaa aattaagggc cttgtgcaac
      301 ccactcgact tcttttggaa tatcttgaag aaaaatatga agagcatttg tatgagcgcg
      361 atgaaggtga taaatggcga aacaaaaagt ttgaattggg tttggagttt cccaatcttc
      421 cttattatat tgatggtgat gttaaattaa cacagtctat ggccatcata cgttatatag
      481 ctgacaagca caacatgttg ggtggttgtc caaaagagcg tgcagagatt tcaatgcttg
      541 aaggagcggt tttggatatt agatacggtg tttcgagaat tgcatatagt aaagactttg
      601 aaactctcaa agttgatttt cttagcaagc tacctgaaat gctgaaaatg ttcgaagatc
      661 gtttatgtca taaaacatat ttaaatggtg atcatgtaac ccatcctgac ttcatgttgt
      721 atgacgctct tgatgttgtt ttatacatgg acccaatgtg cctggatgcg ttcccaaaat
      781 tagtttgttt taaaaaacgt attgaagcta tcccacaaat tgataagtac ttgaaatcca
      841 gcaagtatat agcatggcct ttgcagggct ggcaagccac gtttggtggt ggcgaccatc
      901 ctccaaaatc ggatctggtt ccgcgtggat ccatgagcaa aagcggtgtt ccgcgtctgc
      961 tgaccgcaag cgaacgtgaa cgtctggaac cgtttattga tcagattcat tatagtccgc
     1021 gttatgccga tgatgaatat gaatatcgtc atgtgatgct gccgaaagca atgctgaaag
     1081 caattccgac cgattatttc aatccggaaa ccggtacact gcgtattctg caagaggaag
     1141 aatggcgagg tctgggcatt acccagagct taggttggga aatgtatgaa gttcatgtgc
     1201 cggaaccgca tattctgctg tttaaacgtg aaaaagacta tcagatgaaa ttcagccagc
     1261 agcgtggtgg ttaagcggcc gcatcgtgac tgactgacga tctgcctcgc gcgtttcggt
     1321 gatgacggtg aaaacctctg acacatgcag ctcccggaga cggtcacagc ttgtctgtaa
     1381 gcggatgccg ggagcagaca agcccgtcag ggcgcgtcag cgggtgttgg cgggtgtcgg
     1441 ggcgcagcca tgacccagtc acgtagcgat agcggagtgt ataattcttg aagacgaaag
     1501 ggcctcgtga tacgcctatt tttataggtt aatgtcatga taataatggt ttcttagacg
     1561 tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata
     1621 cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga
     1681 aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca
     1741 ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat
     1801 cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag
     1861 agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc
     1921 gcggtattat cccgtgttga cgccgggcaa gagcaactcg gtcgccgcat acactattct
     1981 cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca
     2041 gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt
     2101 ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat
     2161 gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt
     2221 gacaccacga tgcctgcagc aatggcaaca acgttgcgca aactattaac tggcgaacta
     2281 cttactctag cttcccggca acaattaata gactggatgg aggcggataa agttgcagga
     2341 ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt
     2401 gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc
     2461 gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct
     2521 gagataggtg cctcactgat taagcattgg taactgtcag accaagttta ctcatatata
     2581 ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt
     2641 gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc
     2701 gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg
     2761 caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact
     2821 ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt ccttctagtg
     2881 tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg
     2941 ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac
     3001 tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca
     3061 cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga
     3121 gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc
     3181 ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct
     3241 gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg
     3301 agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct
     3361 tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc
     3421 tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc
     3481 gaggaagcgg aagagcgcct gatgcggtat tttctcctta cgcatctgtg cggtatttca
     3541 caccgcataa attccgacac catcgaatgg cgcaaaacct ttcgcggtat ggcatgatag
     3601 cgcccggaag agagtcaatt cagggtggtg aatgtgaaac cagtaacgtt atacgatgtc
     3661 gcagagtatg ccggtgtctc ttatcagacc gtttcccgcg tggtgaacca ggccagccac
     3721 gtttctgcga aaacgcggga aaaagtggaa gcggcgatgg cggagctgaa ttacattccc
     3781 aaccgcgtgg cacaacaact ggcgggcaaa cagtcgttgc tgattggcgt tgccacctcc
     3841 agtctggccc tgcacgcgcc gtcgcaaatt gtcgcggcga ttaaatctcg cgccgatcaa
     3901 ctgggtgcca gcgtggtggt gtcgatggta gaacgaagcg gcgtcgaagc ctgtaaagcg
     3961 gcggtgcaca atcttctcgc gcaacgcgtc agtgggctga tcattaacta tccgctggat
     4021 gaccaggatg ccattgctgt ggaagctgcc tgcactaatg ttccggcgtt atttcttgat
     4081 gtctctgacc agacacccat caacagtatt attttctccc atgaagacgg tacgcgactg
     4141 ggcgtggagc atctggtcgc attgggtcac cagcaaatcg cgctgttagc gggcccatta
     4201 agttctgtct cggcgcgtct gcgtctggct ggctggcata aatatctcac tcgcaatcaa
     4261 attcagccga tagcggaacg ggaaggcgac tggagtgcca tgtccggttt tcaacaaacc
     4321 atgcaaatgc tgaatgaggg catcgttccc actgcgatgc tggttgccaa cgatcagatg
     4381 gcgctgggcg caatgcgcgc cattaccgag tccgggctgc gcgttggtgc ggatatctcg
     4441 gtagtgggat acgacgatac cgaagacagc tcatgttata tcccgccgtt aaccaccatc
     4501 aaacaggatt ttcgcctgct ggggcaaacc agcgtggacc gcttgctgca actctctcag
     4561 ggccaggcgg tgaagggcaa tcagctgttg cccgtctcac tggtgaaaag aaaaaccacc
     4621 ctggcgccca atacgcaaac cgcctctccc cgcgcgttgg ccgattcatt aatgcagctg
     4681 gcacgacagg tttcccgact ggaaagcggg cagtgagcgc aacgcaatta atgtaagtta
     4741 gctcactcat taggcacccc aggctttaca ctttatgctt ccggctcgta tgttgtgtgg
     4801 aattgtgagc ggataacaat ttcacacagg aaacagctat gaccatgatt acggattcac
     4861 tggccgtcgt tttacaacgt cgtgactggg aaaaccctgg cgttacccaa cttaatcgcc
     4921 ttgcagcaca tccccctttc gccagctggc gtaatagcga agaggcccgc accgatcgcc
     4981 cttcccaaca gttgcgcagc ctgaatggcg aatggcgctt tgcctggttt ccggcaccag
     5041 aagcggtgcc ggaaagctgg ctggagtgcg atcttcctga ggccgatact gtcgtcgtcc
     5101 cctcaaactg gcagatgcac ggttacgatg cgcccatcta caccaacgtg acctatccca
     5161 ttacggtcaa tccgccgttt gttcccacgg agaatccgac gggttgttac tcgctcacat
     5221 ttaatgttga tgaaagctgg ctacaggaag gccagacgcg aattattttt gatggcgttg
     5281 gaattagc
//
