LOCUS       sequence-238928-        7047 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  TRP1-LEU2 ICE plasmid used to facilitate yeast recombination cloning
            and stable genomic Integration after CEN Excision..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7047)
  AUTHORS   Flagg MP, Kao A, Hampton RY
  TITLE     ICE plasmids: combining the ease of yeast recombination cloning with
            the stability of genomic integration.
  JOURNAL   Yeast. 2019 May 10. doi: 10.1002/yea.3400.
  PUBMED    31074531
REFERENCE   2  (bases 1 to 7047)
  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: "Yeast. 2019
            May 10. doi: 10.1002/yea.3400."
COMMENT     Sequence Label: pICE TRP1-LEU2
FEATURES             Location/Qualifiers
     source          1..7047
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(407..1501)
                     /codon_start=1
                     /gene="S. cerevisiae LEU2"
                     /product="3-isopropylmalate dehydrogenase, required for 
                     leucine biosynthesis"
                     /label=LEU2
                     /note="yeast auxotrophic marker"
                     /translation="MSAPKKIVVLPGDHVGQEITAEAIKVLKAISDVRSNVKFDFENHL
                     IGGAAIDATGVPLPDEALEASKKADAVLLGAVGGPKWGTGSVRPEQGLLKIRKELQLYA
                     NLRPCNFASDSLLDLSPIKPQFAKGTDFVVVRELVGGIYFGKRKEDDGDGVAWDSEQYT
                     VPEVQRITRMAAFMALQHEPPLPIWSLDKANVLASSRLWRKTVEETIKNEFPTLKVQHQ
                     LIDSAAMILVKNPTHLNGIIITSNMFGDIISDEASVIPGSLGLLPSASLASLPDKNTAF
                     GLYEPCHGSAPDLPKNKVNPIATILSAAMMLKLSLNLPEEGKAIEDAVKKVLDAGIRTG
                     DLGGSNSTTEVGDAVAEEVKKILA"
     promoter        complement(1502..1909)
                     /gene="S. cerevisiae LEU2"
                     /label=LEU2 promoter
     rep_origin      complement(2210..2665)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2347..2368)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2559..2578
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2791..2813
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2805..2822
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2806..2822
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2832..2851
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2832..2850
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     2875..2891
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     2876..2892
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2926..2942)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2926..2942)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     misc_feature    2973..3476
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     primer_bind     complement(3531..3551)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(3531..3549)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3570..3586)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3570..3586)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3583..3605)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3594..3610
                     /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)."
     promoter        complement(join(3618..3624,3625..3642,3643..3648))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3663..3684
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3801..3818)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3972..4560)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4052..4071)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4731..5522,5523..5591))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     5354..5373
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5592..5696)
                     /gene="bla"
                     /label=AmpR promoter
     promoter        5737..6018
                     /gene="S. cerevisiae TRP1"
                     /label=TRP1 promoter
     CDS             6019..6693
                     /codon_start=1
                     /gene="S. cerevisiae TRP1"
                     /product="phosphoribosylanthranilate isomerase, required 
                     for tryptophan biosynthesis"
                     /label=TRP1
                     /note="yeast auxotrophic marker"
                     /translation="MSVINFTGSSGPLVKVCGLQSTEAAECALDSDADLLGIICVPNRK
                     RTIDPVIARKISSLVKAYKNSSGTPKYLVGVFRNQPKEDVLALVNDYGIDIVQLHGDES
                     WQEYQEFLGLPVIKRLVFPKDCNILLSAASQKPHSFIPLFDSEAGGTGELLDWNSISDW
                     VGRQESPESLHFMLAGGLTPENVGDALRLNGVIGVDVSGGVETNGVKDSNKIANFVKNA
                     KK"
     primer_bind     6763..6781
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(6819..6841)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     6941..6960
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gttcaagaag gtattgactt aaactccatc aaatggtcag gtcattgagt gttttttatt
       61 tgttgtattt tttttttttt agagaaaatc ctccaatata taaattagga atcatagttt
      121 catgattttc tgttacacct aactttttgt gtggtgccct cctccttgtc aatattaatg
      181 ttaaagtgca attctttttc cttatcacgt tgagccatta gtatcaattt gcttacctgt
      241 attcctttac atcctccttt ttctccttct tgataaatgt atgtagattg cgtatatagt
      301 ttcgtctacc ctatgaacat attccatttt gtaatttcgt gtcgtttcta ttatgaattt
      361 catttataaa gtttatgtac aaatatcata aaaaaagaga atctttttaa gcaaggattt
      421 tcttaacttc ttcggcgaca gcatcaccga cttcggtggt actgttggaa ccacctaaat
      481 caccagttct gatacctgca tccaaaacct ttttaactgc atcttcaatg gccttacctt
      541 cttcaggcaa gttcaatgac aatttcaaca tcattgcagc agacaagata gtggcgatag
      601 ggttgacctt attctttggc aaatctggag cagaaccgtg gcatggttcg tacaaaccaa
      661 atgcggtgtt cttgtctggc aaagaggcca aggacgcaga tggcaacaaa cccaaggaac
      721 ctgggataac ggaggcttca tcggagatga tatcaccaaa catgttgctg gtgattataa
      781 taccatttag gtgggttggg ttcttaacta ggatcatggc ggcagaatca atcaattgat
      841 gttgaacctt caatgtaggg aattcgttct tgatggtttc ctccacagtt tttctccata
      901 atcttgaaga ggccaaaaca ttagctttat ccaaggacca aataggcaat ggtggctcat
      961 gttgtagggc catgaaagcg gccattcttg tgattctttg cacttctgga acggtgtatt
     1021 gttcactatc ccaagcgaca ccatcaccat cgtcttcctt tctcttacca aagtaaatac
     1081 ctcccactaa ttctctgaca acaacgaagt cagtaccttt agcaaattgt ggcttgattg
     1141 gagataagtc taaaagagag tcggatgcaa agttacatgg tcttaagttg gcgtacaatt
     1201 gaagttcttt acggattttt agtaaacctt gttcaggtct aacactaccg gtaccccatt
     1261 taggaccacc cacagcacct aacaaaacgg catcagcctt cttggaggct tccagcgcct
     1321 catctggaag tggaacacct gtagcatcga tagcagcacc accaattaaa tgattttcga
     1381 aatcgaactt gacattggaa cgaacatcag aaatagcttt aagaacctta atggcttcgg
     1441 ctgtgatttc ttgaccaacg tggtcacctg gcaaaacgac gatcttctta ggggcagaca
     1501 ttagaatggt atatccttga aatatatata tatatattgc tgaaatgtaa aaggtaagaa
     1561 aagttagaaa gtaagacgat tgctaaccac ctattggaaa aaacaatagg tccttaaata
     1621 atattgtcaa cttcaagtat tgtgatgcaa gcatttagtc atgaacgctt ctctattcta
     1681 tatgaaaagc cggttccggc gctctcacct ttcctttttc tcccaatttt tcagttgaaa
     1741 aaggtatatg cgtcaggcga cctctgaaat taacaaaaaa tttccagtca tcgaatttga
     1801 ttctgtgcga tagcgcccct gtgtgttctc gttatgttga ggaaaaaaat aatggttgct
     1861 aagagattcg aactcttgca tcttacgata cctgagtatt cccacagtta actgcggtca
     1921 agatatttct tgaatcaggc gccttagacc gctcggccaa acaaccaatt acttgttgag
     1981 aaatagagta taattatcct ataaatataa cgtttttgaa cacacatgaa caaggaagta
     2041 caggacaatt gattttgaag agaatgtgga ttttgatgta attgttggga ttccattttt
     2101 aataaggcaa taatattagg tatgtagata tactagaagt tctcctcgac cggtcgatat
     2161 gcggtgtgaa ataccgcaca gatgcgtaag gagaaaatac cgcatcagga aattgtaagc
     2221 gttaatattt tgttaaaatt cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa
     2281 taggccgaaa tcggcaaaat cccttataaa tcaaaagaat agaccgagat agggttgagt
     2341 gttgttccag tttggaacaa gagtccacta ttaaagaacg tggactccaa cgtcaaaggg
     2401 cgaaaaaccg tctatcaggg cgatggccca ctacgtgaac catcacccta atcaagtttt
     2461 ttggggtcga ggtgccgtaa agcactaaat cggaacccta aagggagccc ccgatttaga
     2521 gcttgacggg gaaagccggc gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg
     2581 ggcgctaggg cgctggcaag tgtagcggtc acgctgcgcg taaccaccac acccgccgcg
     2641 cttaatgcgc cgctacaggg cgcgtccatt cgccattcag gctgcgcaac tgttgggaag
     2701 ggcgatcggt gcgggcctct tcgctattac gccagctggc gaaaggggga tgtgctgcaa
     2761 ggcgattaag ttgggtaacg ccagggtttt cccagtcacg acgttgtaaa acgacggcca
     2821 gtgagcgcgc gtaatacgac tcactatagg gcgaattggg taccgggccc cccctcgagg
     2881 tcgacggtat cgataagctt gatatcgaat tcctgcagcc cgggggatcc actagttcta
     2941 gagcggccgc cctcttagcg cttcttagga cggatcgctt gcctgtaact tacacgcgcc
     3001 tcgtatcttt taatgatgga ataatttggg aatttactct gtgtttattt atttttatgt
     3061 tttgtatttg gattttagaa agtaaataaa gaaggtagaa gagttacgga atgaagaaaa
     3121 aaaaataaac aaaggtttaa aaaatttcaa caaaaagcgt actttacata tatatttatt
     3181 agacaagaaa agcagattaa atagatatac attcgattaa cgataagtaa aatgtaaaat
     3241 cacaggattt tcgtgtgtgg tcttctacac agacaagatg aaacaattcg gcattaatac
     3301 ctgagagcag gaagagcaag ataaaaggta gtatttgttg gcgatccccc tagagtcttt
     3361 tacatcttcg gaaaacaaaa actatttttt ctttaatttc tttttttact ttctattttt
     3421 aatttatata tttatattaa aaaatttaaa ttataattat ttttatagca cgtgatgaaa
     3481 aggacccagg tggcgcggcc gccaccgcgg tggagctcca gcttttgttc cctttagtga
     3541 gggttaattg cgcgcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat
     3601 ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc
     3661 taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga
     3721 aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt
     3781 attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg
     3841 cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac
     3901 gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg
     3961 ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca
     4021 agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc
     4081 tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc
     4141 ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag
     4201 gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc
     4261 ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca
     4321 gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg
     4381 aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg
     4441 aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct
     4501 ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa
     4561 gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa
     4621 gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa
     4681 tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc
     4741 ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga
     4801 ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca
     4861 atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc
     4921 ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat
     4981 tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc
     5041 attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt
     5101 tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc
     5161 ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg
     5221 gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt
     5281 gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg
     5341 gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga
     5401 aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg
     5461 taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg
     5521 tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt
     5581 tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc
     5641 atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca
     5701 tttccccgaa aagtgccacc tgacgtctgc accataaacg acattactat atatataata
     5761 taggaagcat ttaatagaac agcatcgtaa tatatgtgta ctttgcagtt atgacgccag
     5821 atggcagtag tggaagatat tctttattga aaaatagctt gtcaccttac gtacaatctt
     5881 gatccggagc ttttcttttt ttgccgatta agaattaatt cggtcgaaaa aagaaaagga
     5941 gagggccaag agggagggca ttggtgacta ttgagcacgt gagtatacgt gattaagcac
     6001 acaaaggcag cttggagtat gtctgttatt aatttcacag gtagttctgg tccattggtg
     6061 aaagtttgcg gcttgcagag cacagaggcc gcagaatgtg ctctagattc cgatgctgac
     6121 ttgctgggta ttatatgtgt gcccaataga aagagaacaa ttgacccggt tattgcaagg
     6181 aaaatttcaa gtcttgtaaa agcatataaa aatagttcag gcactccgaa atacttggtt
     6241 ggcgtgtttc gtaatcaacc taaggaggat gttttggctc tggtcaatga ttacggcatt
     6301 gatatcgtcc aactgcatgg agatgagtcg tggcaagaat accaagagtt cctcggtttg
     6361 ccagttatta aaagactcgt atttccaaaa gactgcaaca tactactcag tgcagcttca
     6421 cagaaacctc attcgtttat tcccttgttt gattcagaag caggtgggac aggtgaactt
     6481 ttggattgga actcgatttc tgactgggtt ggaaggcaag agagccccga aagcttacat
     6541 tttatgttag ctggtggact gacgccagaa aatgttggtg atgcgcttag attaaatggc
     6601 gttattggtg ttgatgtaag cggaggtgtg gagacaaatg gtgtaaaaga ctctaacaaa
     6661 atagcaaatt tcgtcaaaaa tgctaagaaa taggttatta ctgagtagta tttatttaag
     6721 tattgtttgt gcacttgcct aatcatgaca ttaacctata aaaataggcg tatcacgagg
     6781 ccctttcgtc tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg
     6841 gagacggtca cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg
     6901 tcagcgggtg ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta
     6961 ctgagagtgc accatatcga ctacgtcgtt aaggccgttt ctgacagagt aaaattcttg
     7021 agggaacttt caccattatg ggaaatg
//
