LOCUS       sequence-239171-        6458 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  TRP1-HphMx 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 6458)
  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 6458)
  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-HphMx
FEATURES             Location/Qualifiers
     source          1..6458
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(10..32)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     132..151
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        172..453
                     /gene="S. cerevisiae TRP1"
                     /label=TRP1 promoter
     CDS             454..1128
                     /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"
     rep_origin      complement(1225..1680)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1362..1383)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1574..1593
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1806..1828
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     1820..1837
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1821..1837
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1847..1866
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1847..1865
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     1890..1906
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     1891..1907
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1941..1957)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1941..1957)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     misc_feature    1988..2491
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     primer_bind     complement(2546..2566)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(2546..2564)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2585..2601)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2585..2601)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2598..2620)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2609..2625
                     /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(2633..2639,2640..2657,2658..2663))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2678..2699
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2816..2833)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2987..3575)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3067..3086)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3746..4537,4538..4606))
                     /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     4369..4388
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4607..4711)
                     /gene="bla"
                     /label=AmpR promoter
     gene            4784..6359
                     /label=hphMX6
                     /note="yeast selectable marker conferring hygromycin 
                     resistance"
     promoter        4784..5127
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             5128..6156
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     CDS             5128..6156
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     terminator      6162..6359
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     6412..6430
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 cggtgaaaac ctctgacaca tgcagctccc ggagacggtc acagcttgtc tgtaagcgga
       61 tgccgggagc agacaagccc gtcagggcgc gtcagcgggt gttggcgggt gtcggggctg
      121 gcttaactat gcggcatcag agcagattgt actgagagtg caccatagat caacgacatt
      181 actatatata taatatagga agcatttaat agaacagcat cgtaatatat gtgtactttg
      241 cagttatgac gccagatggc agtagtggaa gatattcttt attgaaaaat agcttgtcac
      301 cttacgtaca atcttgatcc ggagcttttc tttttttgcc gattaagaat taattcggtc
      361 gaaaaaagaa aaggagaggg ccaagaggga gggcattggt gactattgag cacgtgagta
      421 tacgtgatta agcacacaaa ggcagcttgg agtatgtctg ttattaattt cacaggtagt
      481 tctggtccat tggtgaaagt ttgcggcttg cagagcacag aggccgcaga atgtgctcta
      541 gattccgatg ctgacttgct gggtattata tgtgtgccca atagaaagag aacaattgac
      601 ccggttattg caaggaaaat ttcaagtctt gtaaaagcat ataaaaatag ttcaggcact
      661 ccgaaatact tggttggcgt gtttcgtaat caacctaagg aggatgtttt ggctctggtc
      721 aatgattacg gcattgatat cgtccaactg catggagatg agtcgtggca agaataccaa
      781 gagttcctcg gtttgccagt tattaaaaga ctcgtatttc caaaagactg caacatacta
      841 ctcagtgcag cttcacagaa acctcattcg tttattccct tgtttgattc agaagcaggt
      901 gggacaggtg aacttttgga ttggaactcg atttctgact gggttggaag gcaagagagc
      961 cccgaaagct tacattttat gttagctggt ggactgacgc cagaaaatgt tggtgatgcg
     1021 cttagattaa atggcgttat tggtgttgat gtaagcggag gtgtggagac aaatggtgta
     1081 aaagactcta acaaaatagc aaatttcgtc aaaaatgcta agaaataggt tattactgag
     1141 tagtatttat ttaattatat cagttattac cctatgcggt gtgaaatacc gcacagatgc
     1201 gtaaggagaa aataccgcat caggaaattg taagcgttaa tattttgtta aaattcgcgt
     1261 taaatttttg ttaaatcagc tcatttttta accaataggc cgaaatcggc aaaatccctt
     1321 ataaatcaaa agaatagacc gagatagggt tgagtgttgt tccagtttgg aacaagagtc
     1381 cactattaaa gaacgtggac tccaacgtca aagggcgaaa aaccgtctat cagggcgatg
     1441 gcccactacg tgaaccatca ccctaatcaa gttttttggg gtcgaggtgc cgtaaagcac
     1501 taaatcggaa ccctaaaggg agcccccgat ttagagcttg acggggaaag ccggcgaacg
     1561 tggcgagaaa ggaagggaag aaagcgaaag gagcgggcgc tagggcgctg gcaagtgtag
     1621 cggtcacgct gcgcgtaacc accacacccg ccgcgcttaa tgcgccgcta cagggcgcgt
     1681 ccattcgcca ttcaggctgc gcaactgttg ggaagggcga tcggtgcggg cctcttcgct
     1741 attacgccag ctggcgaaag ggggatgtgc tgcaaggcga ttaagttggg taacgccagg
     1801 gttttcccag tcacgacgtt gtaaaacgac ggccagtgag cgcgcgtaat acgactcact
     1861 atagggcgaa ttgggtaccg ggccccccct cgaggtcgac ggtatcgata agcttgatat
     1921 cgaattcctg cagcccgggg gatccactag ttctagagcg gccgccctct tagcgcttct
     1981 taggacggat cgcttgcctg taacttacac gcgcctcgta tcttttaatg atggaataat
     2041 ttgggaattt actctgtgtt tatttatttt tatgttttgt atttggattt tagaaagtaa
     2101 ataaagaagg tagaagagtt acggaatgaa gaaaaaaaaa taaacaaagg tttaaaaaat
     2161 ttcaacaaaa agcgtacttt acatatatat ttattagaca agaaaagcag attaaataga
     2221 tatacattcg attaacgata agtaaaatgt aaaatcacag gattttcgtg tgtggtcttc
     2281 tacacagaca agatgaaaca attcggcatt aatacctgag agcaggaaga gcaagataaa
     2341 aggtagtatt tgttggcgat ccccctagag tcttttacat cttcggaaaa caaaaactat
     2401 tttttcttta atttcttttt ttactttcta tttttaattt atatatttat attaaaaaat
     2461 ttaaattata attattttta tagcacgtga tgaaaaggac ccaggtggcg cggccgccac
     2521 cgcggtggag ctccagcttt tgttcccttt agtgagggtt aattgcgcgc ttggcgtaat
     2581 catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac
     2641 gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa
     2701 ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat
     2761 gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc
     2821 tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg
     2881 cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag
     2941 gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc
     3001 gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag
     3061 gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga
     3121 ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc
     3181 atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg
     3241 tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt
     3301 ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca
     3361 gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca
     3421 ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag
     3481 ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca
     3541 agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg
     3601 ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa
     3661 aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta
     3721 tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag
     3781 cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga
     3841 tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac
     3901 cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc
     3961 ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta
     4021 gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac
     4081 gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat
     4141 gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa
     4201 gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg
     4261 tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag
     4321 aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc
     4381 cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct
     4441 caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat
     4501 cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg
     4561 ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc
     4621 aatattattg aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta
     4681 tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg
     4741 tcttgtttag cttgcctcgt ccccgccggg tcacccggcc agcgacatgg aggcccagaa
     4801 taccctcctt gacagtcttg acgtgcgcag ctcaggggca tgatgtgact gtcgcccgta
     4861 catttagccc atacatcccc atgtataatc atttgcatcc atacattttg atggccgcac
     4921 ggcgcgaagc aaaaattacg gctcctcgct gcggacctgc gagcagggaa acgctcccct
     4981 cacagacgcg ttgaattgtc cccacgccgc gcccctgtag agaaatataa aaggttagga
     5041 tttgccactg aggttcttct ttcatatact tccttttaaa atcttgctag gatacagttc
     5101 tcacatcaca tccgaacata aacaaccatg ggtaaaaagc ctgaactcac cgcgacgtct
     5161 gtcgagaagt ttctgatcga aaagttcgac agcgtctccg acctgatgca gctctcggag
     5221 ggcgaagaat ctcgtgcttt cagcttcgat gtaggagggc gtggatatgt cctgcgggta
     5281 aatagctgcg ccgatggttt ctacaaagat cgttatgttt atcggcactt tgcatcggcc
     5341 gcgctcccga ttccggaagt gcttgacatt ggggaattca gcgagagcct gacctattgc
     5401 atctcccgcc gtgcacaggg tgtcacgttg caagacctgc ctgaaaccga actgcccgct
     5461 gttctgcagc cggtcgcgga ggccatggat gcgatcgctg cggccgatct tagccagacg
     5521 agcgggttcg gcccattcgg accgcaagga atcggtcaat acactacatg gcgtgatttc
     5581 atatgcgcga ttgctgatcc ccatgtgtat cactggcaaa ctgtgatgga cgacaccgtc
     5641 agtgcgtccg tcgcgcaggc tctcgatgag ctgatgcttt gggccgagga ctgccccgaa
     5701 gtccggcacc tcgtgcacgc ggatttcggc tccaacaatg tcctgacgga caatggccgc
     5761 ataacagcgg tcattgactg gagcgaggcg atgttcgggg attcccaata cgaggtcgcc
     5821 aacatcttct tctggaggcc gtggttggct tgtatggagc agcagacgcg ctacttcgag
     5881 cggaggcatc cggagcttgc aggatcgccg cggctccggg cgtatatgct ccgcattggt
     5941 cttgaccaac tctatcagag cttggttgac ggcaatttcg atgatgcagc ttgggcgcag
     6001 ggtcgatgcg acgcaatcgt ccgatccgga gccgggactg tcgggcgtac acaaatcgcc
     6061 cgcagaagcg cggccgtctg gaccgatggc tgtgtagaag tactcgccga tagtggaaac
     6121 cgacgcccca gcactcgtcc gagggcaaag gaataatcag tactgacaat aaaaagattc
     6181 ttgttttcaa gaacttgtca tttgtatagt ttttttatat tgtagttgtt ctattttaat
     6241 caaatgttag cgtgatttat attttttttc gcctcgacat catctgccca gatgcgaagt
     6301 taagtgcgca gaaagtaata tcatgcgtca atcgtatgtg aatgctggtc gctatactgc
     6361 tgtcgattcg atactaacgc cgccatccag atcatgacat taacctataa aaataggcgt
     6421 atcacgaggc cctttcgtct cgcgcgtttc ggtgatga
//
