LOCUS       sequence-10375-e        5512 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5512)
  AUTHORS   Miyake S, Sellers WR, Safran M, Li X, Zhao W, Grossman SR, Gan J, 
            DeCaprio JA, Adams PD, Kaelin WG
  TITLE     Cells degrade a novel inhibitor of differentiation with E1A-like 
            properties upon exiting the cell cycle.
  JOURNAL   Mol Cell Biol. 2000 Dec . 20(23):8889-902.
  PUBMED    11073989
REFERENCE   2  (bases 1 to 5512)
  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: "Mol Cell 
            Biol. 2000 Dec . 20(23):8889-902."
COMMENT     Sequence Label: EID1-pGex-2TK
FEATURES             Location/Qualifiers
     source          1..5512
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(183..188,189..204,205..211)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    219..235
                     /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             258..911
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     869..891
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             918..935
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     primer_bind     complement(1562..1584)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1729..1747)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1815..1919
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1920..1988,1989..2780)
                     /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(2138..2157)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2951..3539
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3440..3459
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3693..3710
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        3783..3860
                     /gene="lacI (mutant)"
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the 
                     promoter boosts expression of the lacI gene about 10-fold."
     CDS             3861..4943
                     /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(3880..3899)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     protein_bind    4956..4977
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(4992..4997,4998..5015,5016..5022)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5030..5046
                     /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     5054..5070
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5054..5070
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             join(5066..5512,1..81)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGITLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(5086..5103)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5086..5102)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5095..5117)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 acgttatcga ctgcacggtg caccaatgct tctggcgtca ggcagccatc ggaagctgtg
       61 gtatggctgt gcaggtcgta aatcactgca taattcgtgt cgctcaaggc gcactcccgt
      121 tctggataat gttttttgcg ccgacatcat aacggttctg gcaaatattc tgaaatgagc
      181 tgttgacaat taatcatcgg ctcgtataat gtgtggaatt gtgagcggat aacaatttca
      241 cacaggaaac agtattcatg tcccctatac taggttattg gaaaattaag ggccttgtgc
      301 aacccactcg acttcttttg gaatatcttg aagaaaaata tgaagagcat ttgtatgagc
      361 gcgatgaagg tgataaatgg cgaaacaaaa agtttgaatt gggtttggag tttcccaatc
      421 ttccttatta tattgatggt gatgttaaat taacacagtc tatggccatc atacgttata
      481 tagctgacaa gcacaacatg ttgggtggtt gtccaaaaga gcgtgcagag atttcaatgc
      541 ttgaaggagc ggttttggat attagatacg gtgtttcgag aattgcatat agtaaagact
      601 ttgaaactct caaagttgat tttcttagca agctacctga aatgctgaaa atgttcgaag
      661 atcgtttatg tcataaaaca tatttaaatg gtgatcatgt aacccatcct gacttcatgt
      721 tgtatgacgc tcttgatgtt gttttataca tggacccaat gtgcctggat gcgttcccaa
      781 aattagtttg ttttaaaaaa cgtattgaag ctatcccaca aattgataag tacttgaaat
      841 ccagcaagta tatagcatgg cctttgcagg gctggcaagc cacgtttggt ggtggcgacc
      901 atcctccaaa atcggatctg gttccgcgtg gatccgccac catgtcggaa atggctgagt
      961 tgtccgagct gtatgaagag agcagtgacc tgcagatgga tgtgatgcct ggcgagggtg
     1021 accttccgca gatggaggta ggcagcggga gccgggagct atccctgcgt ccctcccgca
     1081 gcggggccca acagctcgag gaggaaggcc caatggagga ggaggaggcc cagccaatgg
     1141 cggcgccaga ggggaaacgg agccttgcta acgggcccaa cgctggggag cagccaggcc
     1201 aggtggcggg cgcagacttc gagagcgagg acgagggcga ggaatttgat gactgggagg
     1261 acgactacga ctatcccgaa gaggagcagc tcagtggtgc cggctacaga gtatcagccg
     1321 ctcttgaaga agccgacaag atgtttctga gaacaagaga accagccctg gatggcgggt
     1381 ttcagatgca ttatgagaag accccgtttg atcagttagc ttttatcgaa gagctttttt
     1441 cactgatggt tgtcaatcgt ctgaccgaag aactcggctg tgatgagatt attgatagag
     1501 agtgaattca tcgtgactga ctgacgatct gcctcgcgcg tttcggtgat gacggtgaaa
     1561 acctctgaca catgcagctc ccggagacgg tcacagcttg tctgtaagcg gatgccggga
     1621 gcagacaagc ccgtcagggc gcgtcagcgg gtgttggcgg gtgtcggggc gcagccatga
     1681 cccagtcacg tagcgatagc ggagtgtata attcttgaag acgaaagggc ctcgtgatac
     1741 gcctattttt ataggttaat gtcatgataa taatggtttc ttagacgtca ggtggcactt
     1801 ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt
     1861 atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta
     1921 tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg
     1981 tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac
     2041 gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg
     2101 aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc
     2161 gtgttgacgc cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg
     2221 ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat
     2281 gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg
     2341 gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg
     2401 atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc
     2461 ctgcagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt
     2521 cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct
     2581 cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc
     2641 gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca
     2701 cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct
     2761 cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt
     2821 taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga
     2881 ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca
     2941 aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac
     3001 caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg
     3061 taactggctt cagcagagcg cagataccaa atactgtcct tctagtgtag ccgtagttag
     3121 gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac
     3181 cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt
     3241 taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg
     3301 agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc
     3361 ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc
     3421 gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc
     3481 acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa
     3541 acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt
     3601 tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg
     3661 ataccgctcg ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag
     3721 agcgcctgat gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcataaatt
     3781 ccgacaccat cgaatggtgc aaaacctttc gcggtatggc atgatagcgc ccggaagaga
     3841 gtcaattcag ggtggtgaat gtgaaaccag taacgttata cgatgtcgca gagtatgccg
     3901 gtgtctctta tcagaccgtt tcccgcgtgg tgaaccaggc cagccacgtt tctgcgaaaa
     3961 cgcgggaaaa agtggaagcg gcgatggcgg agctgaatta cattcccaac cgcgtggcac
     4021 aacaactggc gggcaaacag tcgttgctga ttggcgttgc cacctccagt ctggccctgc
     4081 acgcgccgtc gcaaattgtc gcggcgatta aatctcgcgc cgatcaactg ggtgccagcg
     4141 tggtggtgtc gatggtagaa cgaagcggcg tcgaagcctg taaagcggcg gtgcacaatc
     4201 ttctcgcgca acgcgtcagt gggctgatca ttaactatcc gctggatgac caggatgcca
     4261 ttgctgtgga agctgcctgc actaatgttc cggcgttatt tcttgatgtc tctgaccaga
     4321 cacccatcaa cagtattatt ttctcccatg aagacggtac gcgactgggc gtggagcatc
     4381 tggtcgcatt gggtcaccag caaatcgcgc tgttagcggg cccattaagt tctgtctcgg
     4441 cgcgtctgcg tctggctggc tggcataaat atctcactcg caatcaaatt cagccgatag
     4501 cggaacggga aggcgactgg agtgccatgt ccggttttca acaaaccatg caaatgctga
     4561 atgagggcat cgttcccact gcgatgctgg ttgccaacga tcagatggcg ctgggcgcaa
     4621 tgcgcgccat taccgagtcc gggctgcgcg ttggtgcgga tatctcggta gtgggatacg
     4681 acgataccga agacagctca tgttatatcc cgccgttaac caccatcaaa caggattttc
     4741 gcctgctggg gcaaaccagc gtggaccgct tgctgcaact ctctcagggc caggcggtga
     4801 agggcaatca gctgttgccc gtctcactgg tgaaaagaaa aaccaccctg gcgcccaata
     4861 cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca cgacaggttt
     4921 cccgactgga aagcgggcag tgagcgcaac gcaattaatg tgagttagct cactcattag
     4981 gcaccccagg ctttacactt tatgcttccg gctcgtatgt tgtgtggaat tgtgagcgga
     5041 taacaatttc acacaggaaa cagctatgac catgattacg gattcactgg ccgtcgtttt
     5101 acaacgtcgt gactgggaaa accctggcgt tacccaactt aatcgccttg cagcacatcc
     5161 ccctttcgcc agctggcgta atagcgaaga ggcccgcacc gatcgccctt cccaacagtt
     5221 gcgcagcctg aatggcgaat ggcgctttgc ctggtttccg gcaccagaag cggtgccgga
     5281 aagctggctg gagtgcgatc ttcctgaggc cgatactgtc gtcgtcccct caaactggca
     5341 gatgcacggt tacgatgcgc ccatctacac caacgtaacc tatcccatta cggtcaatcc
     5401 gccgtttgtt cccacggaga atccgacggg ttgttactcg ctcacattta atgttgatga
     5461 aagctggcta caggaaggcc agacgcgaat tatttttgat ggcgttggaa tt
//
