LOCUS       sequence-234447-        6006 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  TRP1-NatMx 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 6006)
  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 6006)
  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-NatMx
FEATURES             Location/Qualifiers
     source          1..6006
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(29..51)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     151..170
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        187..467
                     /gene="S. cerevisiae TRP1"
                     /label=TRP1 promoter
     CDS             468..1142
                     /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(1241..1696)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1378..1399)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1590..1609
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1826..1848
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     1840..1857
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1841..1857
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1867..1886
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1867..1885
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     1910..1926
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     1911..1927
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1961..1977)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1961..1977)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     misc_feature    2008..2511
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     primer_bind     complement(2566..2586)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(2566..2584)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2605..2621)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2605..2621)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2618..2640)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2629..2645
                     /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(2653..2659,2660..2677,2678..2683))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     primer_bind     complement(2836..2853)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3007..3595)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3087..3106)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3766..4557,4558..4626))
                     /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     4389..4408
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4627..4731)
                     /gene="bla"
                     /label=AmpR promoter
     gene            4804..5926
                     /label=natMX6
                     /note="yeast selectable marker conferring nourseothricin 
                     resistance"
     promoter        4804..5147
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             5148..5723
                     /codon_start=1
                     /gene="Streptomyces noursei nat1"
                     /product="nourseothricin acetyltransferase"
                     /label=NrsR
                     /note="confers resistance to nourseothricin"
                     /translation="MGTTLDDTAYRYRTSVPGDAEAIEALDGSFTTDTVFRVTATGDGF
                     TLREVPVDPPLTKVFPDDESDDESDDGEDGDPDSRTFVAYGDDGDLAGFVVISYSAWNR
                     RLTVEDIEVAPEHRGHGVGRALMGLATEFAGERGAGHLWLEVTNVNAPAIHAYRRMGFT
                     LCGLDTALYDGTASDGERQALYMSMPCP"
     terminator      5729..5926
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     5979..5997
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
      181 accataaacg acattactat atatataata taggaagcat ttaatagaca gcatcgtaat
      241 atatgtgtac tttgcagtta tgacgccaga tggcagtagt ggaagatatt ctttattgaa
      301 aaatagcttg tcaccttacg tacaatcttg atccggagct tttctttttt tgccgattaa
      361 gaattaattc ggtcgaaaaa agaaaaggag agggccaaga gggagggcat tggtgactat
      421 tgagcacgtg agtatacgtg attaagcaca caaaggcagc ttggagtatg tctgttatta
      481 atttcacagg tagttctggt ccattggtga aagtttgcgg cttgcagagc acagaggccg
      541 cagaatgtgc tctagattcc gatgctgact tgctgggtat tatatgtgtg cccaatagaa
      601 agagaacaat tgacccggtt attgcaagga aaatttcaag tcttgtaaaa gcatataaaa
      661 atagttcagg cactccgaaa tacttggttg gcgtgtttcg taatcaacct aaggaggatg
      721 ttttggctct ggtcaatgat tacggcattg atatcgtcca actgcatgga gatgagtcgt
      781 ggcaagaata ccaagagttc ctcggtttgc cagttattaa aagactcgta tttccaaaag
      841 actgcaacat actactcagt gcagcttcac agaaacctca ttcgtttatt cccttgtttg
      901 attcagaagc aggtgggaca ggtgaacttt tggattggaa ctcgatttct gactgggttg
      961 gaaggcaaga gagccccgaa agcttacatt ttatgttagc tggtggactg acgccagaaa
     1021 atgttggtga tgcgcttaga ttaaatggcg ttattggtgt tgatgtaagc ggaggtgtgg
     1081 agacaaatgg tgtaaaagac tctaacaaaa tagcaaattt cgtcaaaaat gctaagaaat
     1141 aggttattac tgagtagtat ttatttaagt attgtttgtg cacttgccta tgcggtgtga
     1201 aataccgcac agatgcgtaa ggagaaaata ccgcatcagg aaattgtaaa cgttaatatt
     1261 ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat tttttaacca ataggccgaa
     1321 atcggcaaaa tcccttataa atcaaaagaa tagaccgaga tagggttgag tgttgttcca
     1381 gtttggaaca agagtccact attaaagaac gtggactcca acgtcaaagg gcgaaaaacc
     1441 gtctatcagg gcgatggccc actacgtgaa ccatcaccct aatcaagttt tttggggtcg
     1501 aggtgccgta aagcactaaa tcggaaccct aaagggagcc cccgatttag agcttgacgg
     1561 ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag cgaaaggagc gggcgctagg
     1621 gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca cacccgccgc gcttaatgcg
     1681 ccgctacagg gcgcgtcgcg ccattcgcca ttcaggctgc gcaactgttg ggaagggcga
     1741 tcggtgcggg cctcttcgct attacgccag ctggcgaaag ggggatgtgc tgcaaggcga
     1801 ttaagttggg taacgccagg gttttcccag tcacgacgtt gtaaaacgac ggccagtgag
     1861 cgcgcgtaat acgactcact atagggcgaa ttgggtaccg ggccccccct cgaggtcgac
     1921 ggtatcgata agcttgatat cgaattcctg cagcccgggg gatccactag ttctagagcg
     1981 gccgccctct tagcgcttct taggacggat cgcttgcctg taacttacac gcgcctcgta
     2041 tcttttaatg atggaataat ttgggaattt actctgtgtt tatttatttt tatgttttgt
     2101 atttggattt tagaaagtaa ataaagaagg tagaagagtt acggaatgaa gaaaaaaaaa
     2161 taaacaaagg tttaaaaaat ttcaacaaaa agcgtacttt acatatatat ttattagaca
     2221 agaaaagcag attaaataga tatacattcg attaacgata agtaaaatgt aaaatcacag
     2281 gattttcgtg tgtggtcttc tacacagaca agatgaaaca attcggcatt aatacctgag
     2341 agcaggaaga gcaagataaa aggtagtatt tgttggcgat ccccctagag tcttttacat
     2401 cttcggaaaa caaaaactat tttttcttta atttcttttt ttactttcta tttttaattt
     2461 atatatttat attaaaaaat ttaaattata attattttta tagcacgtga tgaaaaggac
     2521 ccaggtggcg cggccgccac cgcggtggag ctccagcttt tgttcccttt agtgagggtt
     2581 aattgcgcgc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt gttatccgct
     2641 cacaattcca cacaacatag gagccggaag cataaagtgt aaagcctggg gtgcctaatg
     2701 agtgaggtaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct
     2761 gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg
     2821 gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc
     2881 ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg
     2941 aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct
     3001 ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca
     3061 gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct
     3121 cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc
     3181 gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt
     3241 tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc
     3301 cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc
     3361 cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg
     3421 gtggcctaac tacggctaca ctagaaggac agtatttggt atctgcgctc tgctgaagcc
     3481 agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag
     3541 cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga
     3601 tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat
     3661 tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag
     3721 ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat
     3781 cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc
     3841 cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat
     3901 accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag
     3961 ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg
     4021 ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc
     4081 tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca
     4141 acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg
     4201 tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc
     4261 actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta
     4321 ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc
     4381 aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg
     4441 ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc
     4501 cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc
     4561 aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat
     4621 actcatactc ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag
     4681 cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc
     4741 ccgaaaagtg ccacctgacg tcttgtttag cttgcctcgt ccccgccggg tcacccggcc
     4801 agcgacatgg aggcccagaa taccctcctt gacagtcttg acgtgcgcag ctcaggggca
     4861 tgatgtgact gtcgcccgta catttagccc atacatcccc atgtataatc atttgcatcc
     4921 atacattttg atggccgcac ggcgcgaagc aaaaattacg gctcctcgct gcagacctgc
     4981 gagcagggaa acgctcccct cacagacgcg ttgaattgtc cccacgccgc gcccctgtag
     5041 agaaatataa aaggttagga tttgccactg aggttcttct ttcatatact tccttttaaa
     5101 atcttgctag gatacagttc tcacatcaca tccgaacata aacaaccatg ggtaccactc
     5161 ttgacgacac ggcttaccgg taccgcacca gtgtcccggg ggacgccgag gccatcgagg
     5221 cactggatgg gtccttcacc accgacaccg tcttccgcgt caccgccacc ggggacggct
     5281 tcaccctgcg ggaggtgccg gtggacccgc ccctgaccaa ggtgttcccc gacgacgaat
     5341 cggacgacga atcggacgac ggggaggacg gcgacccgga ctcccggacg ttcgtcgcgt
     5401 acggggacga cggcgacctg gcgggcttcg tggtcatctc gtactcggcg tggaaccgcc
     5461 ggctgaccgt cgaggacatc gaggtcgccc cggagcaccg ggggcacggg gtcgggcgcg
     5521 cgttgatggg gctcgcgacg gagttcgccg gcgagcgggg cgccgggcac ctctggctgg
     5581 aggtcaccaa cgtcaacgca ccggcgatcc acgcgtaccg gcggatgggg ttcaccctct
     5641 gcggcctgga caccgccctg tacgacggca ccgcctcgga cggcgagcgg caggcgctct
     5701 acatgagcat gccctgcccc taatcagtac tgacaataaa aagattcttg ttttcaagaa
     5761 cttgtcattt gtatagtttt tttatattgt agttgttcta ttttaatcaa atgttagcgt
     5821 gatttatatt ttttttcgcc tcgacatcat ctgcccagat gcgaagttaa gtgcgcagaa
     5881 agtaatatca tgcgtcaatc gtatgtgaat gctggtcgct atactgctgt cgattcgata
     5941 ctaacgccgc catccagatc atgacattaa cctataaaaa taggcgtatc acgaggccct
     6001 ttcgtc
//
