LOCUS       sequence-239191-        7415 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  ADE2-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 7415)
  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 7415)
  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 ADE2-NatMx
FEATURES             Location/Qualifiers
     source          1..7415
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     73..92
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        283..658
                     /gene="S. cerevisiae ADE2"
                     /label=ADE2 promoter
     CDS             659..2374
                     /codon_start=1
                     /gene="S. cerevisiae ADE2"
                     /product="phosphoribosylaminoimidazole carboxylase, 
                     required for adenine biosynthesis"
                     /label=ADE2
                     /note="yeast auxotrophic marker"
                     /translation="MDSRTVGILGGGQLGRMIVEAANRLNIKTVILDAENSPAKQISNS
                     NDHVNGSFSNPLDIEKLAEKCDVLTIEIEHVDVPTLKNLQVKHPKLKIYPSPETIRLIQ
                     DKYIQKEHLIKNGIAVTQSVPVEQASETSLLNVGRDLGFPFVLKSRTLAYDGRGNFVVK
                     NKEMIPEALEVLKDRPLYAEKWAPFTKELAVMIVRSVNGLVFSYPIVETIHKDNICDLC
                     YAPARVPDSVQLKAKLLAENAIKSFPGCGIFGVEMFYLETGELLINEIAPRPHNSGHYT
                     IDACVTSQFEAHLRSILDLPMPKNFTSFSTITTNAIMLNVLGDKHTKDKELETCERALA
                     TPGSSVYLYGKESRPNRKVGHINIIASSMAECEQRLNYITGRTDIPIKISVAQKLDLEA
                     MVKPLVGIIMGSDSDLPVMSAACAVLKDFGVPFEVTIVSAHRTPHRMSAYAISASKRGI
                     KTIIAGAGGAAHLPGMVAAMTPLPVIGVPVKGSCLDGVDSLHSIVQMPRGVPVATVAIN
                     NSTNAALLAVRLLGAYDSSYTTKMEQFLLKQEEEVLVKAQKLETVGYEAYLENK"
     rep_origin      complement(2579..3034)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2716..2737)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2928..2947
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3160..3182
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     3174..3191
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     3175..3191
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3201..3220
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3201..3219
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     3244..3260
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     3245..3261
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3295..3311)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3295..3311)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     misc_feature    3342..3845
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     primer_bind     complement(3900..3920)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(3900..3918)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3939..3955)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3939..3955)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3952..3974)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3963..3979
                     /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(3987..3993,3994..4011,4012..4017))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4032..4053
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4170..4187)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4341..4929)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4421..4440)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5100..5891,5892..5960))
                     /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     5723..5742
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5961..6065)
                     /gene="bla"
                     /label=AmpR promoter
     gene            6138..7257
                     /label=natMX6
                     /note="yeast selectable marker conferring nourseothricin 
                     resistance"
     promoter        6138..6481
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             6482..7054
                     /codon_start=1
                     /gene="Streptomyces noursei nat1"
                     /product="nourseothricin acetyltransferase"
                     /label=NrsR
                     /note="confers resistance to nourseothricin"
                     /translation="MGTTLDDTAYRYRTSVPGDAEAIEALDGSFTTDTVFRVTATGDGF
                     TLREVPVDPPLTKVFPDDESDDESDDGEDGDPDSRTFVAYGDDGDLAGFVVVSYSGWNR
                     RLTVEDIEVAPEHRGHGVGRALMGLATEFARERGAGHLWLEVTNVNAPAIHAYRRMGFT
                     LCGLDTALYDGTASDGEQALYMSMPCP"
     terminator      7060..7257
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     7310..7328
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(7366..7388)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
ORIGIN
        1 atgccgggag cagacaagcc cgtcagggcg cgtcagcggg tgttggcggg tgtcggggct
       61 ggcttaacta tgcggcatca gagcagattg tactgagagt gcaccatatg cggtgtgaaa
      121 taccgcacag atgcgtaagg agaaaatacc gcatcaggaa attgtaagcg ttaatatttt
      181 gttaaaattc gcgttaaatt tttgttaaat cagctcattt tttaaccaat aggccgaaat
      241 cggcaaaatc ccttataaat caaaagaata gatctgaatt aattcttgaa taatacataa
      301 cttttcttaa aagaatcaaa gacagataaa atttaagaga tattaaatat tagtgagaag
      361 ccgagaattt tgtaacacca acataacact gacatcttta acaactttta attatgatac
      421 atttcttacg tcatgattga ttattacagc tatgctgaca aatgactctt gttgcatggc
      481 tacgaaccgg gtaatactaa gtgattgact cttgctgacc ttttattaag aactaaatgg
      541 acaatattat ggagcatttc atgtataaat tggtgcgtaa aatcgttgga tctctcttct
      601 aagtacatcc tactataaca atcaagaaaa acaagaaaat cggacaaaac aatcaagtat
      661 ggattctaga acagttggta tattaggagg gggacaattg ggacgtatga ttgttgaggc
      721 agcaaacagg ctcaacatta agacggtaat actagatgct gaaaattctc ctgccaaaca
      781 aataagcaac tccaatgacc acgttaatgg ctccttttcc aatcctcttg atatcgaaaa
      841 actagctgaa aaatgtgatg tgctaacgat tgagattgag catgttgatg ttcctacact
      901 aaagaatctt caagtaaaac atcccaaatt aaaaatttac ccttctccag aaacaatcag
      961 attgatacaa gacaaatata ttcaaaaaga gcatttaatc aaaaatggta tagcagttac
     1021 ccaaagtgtt cctgtggaac aagccagtga gacgtcccta ttgaatgttg gaagagattt
     1081 gggttttcca ttcgtcttga agtcgaggac tttggcatac gatggaagag gtaacttcgt
     1141 tgtaaagaat aaggaaatga ttccggaagc tttggaagta ctgaaggatc gtcctttgta
     1201 cgccgaaaaa tgggcaccat ttactaaaga attagcagtc atgattgtga ggtctgttaa
     1261 cggtttagtg ttttcttacc caattgtaga gactatccac aaggacaata tttgtgactt
     1321 atgttatgcg cctgctagag ttccggactc cgttcaactt aaggcgaagt tgttggcaga
     1381 aaatgcaatc aaatcttttc ccggttgtgg tatatttggt gtggaaatgt tctatttaga
     1441 aacaggggaa ttgcttatta acgaaattgc cccaaggcct cacaactctg gacattatac
     1501 cattgatgct tgcgtcactt ctcaatttga agctcatttg agatcaatat tggatttgcc
     1561 aatgccaaag aatttcacat ctttctccac cattacaacg aacgccatta tgctaaatgt
     1621 tcttggagac aaacatacaa aagataaaga gctagaaact tgcgaaagag cattggcgac
     1681 tccaggttcc tcagtgtact tatatggaaa agagtctaga cctaacagaa aagtaggtca
     1741 cataaatatt attgcctcca gtatggcgga atgtgaacaa aggctgaact acattacagg
     1801 tagaactgat attccaatca aaatctctgt cgctcaaaag ttggacttgg aagcaatggt
     1861 caaaccattg gttggaatca tcatgggatc agactctgac ttgccggtaa tgtctgccgc
     1921 atgtgcggtt ttaaaagatt ttggcgttcc atttgaagtg acaatagtct ctgctcatag
     1981 aactccacat aggatgtcag catatgctat ttccgcaagc aagcgtggaa ttaaaacaat
     2041 tatcgctgga gctggtgggg ctgctcactt gccaggtatg gtggctgcaa tgacaccact
     2101 tcctgtcatc ggtgtgcccg taaaaggttc ttgtctagat ggagtagatt ctttacattc
     2161 aattgtgcaa atgcctagag gtgttccagt agctaccgtc gctattaata atagtacgaa
     2221 cgctgcgctg ttggctgtca gactgcttgg cgcttatgat tcaagttata caacgaaaat
     2281 ggaacagttt ttattaaagc aagaagaaga agttcttgtc aaagcacaaa agttagaaac
     2341 tgtcggttac gaagcttatc tagaaaacaa gtaatatata agtttattga tatacttgta
     2401 cagcaaataa ttataaaatg atatacctat tttttaggct ttgttatgat tacatcaaat
     2461 gtggacttca tacatagaaa tcaacgctta caggtgtcct tttttaagaa tttcatacat
     2521 aagatctatg cggtgtgaaa taccgcacag atgcgtaagg agaaaatacc gcatcaggaa
     2581 attgtaagcg ttaatatttt gttaaaattc gcgttaaatt tttgttaaat cagctcattt
     2641 tttaaccaat aggccgaaat cggcaaaatc ccttataaat caaaagaata gaccgagata
     2701 gggttgagtg ttgttccagt ttggaacaag agtccactat taaagaacgt ggactccaac
     2761 gtcaaagggc gaaaaaccgt ctatcagggc gatggcccac tacgtgaacc atcaccctaa
     2821 tcaagttttt tggggtcgag gtgccgtaaa gcactaaatc ggaaccctaa agggagcccc
     2881 cgatttagag cttgacgggg aaagccggcg aacgtggcga gaaaggaagg gaagaaagcg
     2941 aaaggagcgg gcgctagggc gctggcaagt gtagcggtca cgctgcgcgt aaccaccaca
     3001 cccgccgcgc ttaatgcgcc gctacagggc gcgtccattc gccattcagg ctgcgcaact
     3061 gttgggaagg gcgatcggtg cgggcctctt cgctattacg ccagctggcg aaagggggat
     3121 gtgctgcaag gcgattaagt tgggtaacgc cagggttttc ccagtcacga cgttgtaaaa
     3181 cgacggccag tgagcgcgcg taatacgact cactataggg cgaattgggt accgggcccc
     3241 ccctcgaggt cgacggtatc gataagcttg atatcgaatt cctgcagccc gggggatcca
     3301 ctagttctag agcggccgcc ctcttagcgc ttcttaggac ggatcgcttg cctgtaactt
     3361 acacgcgcct cgtatctttt aatgatggaa taatttggga atttactctg tgtttattta
     3421 tttttatgtt ttgtatttgg attttagaaa gtaaataaag aaggtagaag agttacggaa
     3481 tgaagaaaaa aaaataaaca aaggtttaaa aaatttcaac aaaaagcgta ctttacatat
     3541 atatttatta gacaagaaaa gcagattaaa tagatataca ttcgattaac gataagtaaa
     3601 atgtaaaatc acaggatttt cgtgtgtggt cttctacaca gacaagatga aacaattcgg
     3661 cattaatacc tgagagcagg aagagcaaga taaaaggtag tatttgttgg cgatccccct
     3721 agagtctttt acatcttcgg aaaacaaaaa ctattttttc tttaatttct ttttttactt
     3781 tctattttta atttatatat ttatattaaa aaatttaaat tataattatt tttatagcac
     3841 gtgatgaaaa ggacccaggt ggcgcggccg ccaccgcggt ggagctccag cttttgttcc
     3901 ctttagtgag ggttaattgc gcgcttggcg taatcatggt catagctgtt tcctgtgtga
     3961 aattgttatc cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc
     4021 tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc
     4081 cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc
     4141 ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt
     4201 cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca
     4261 ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa
     4321 aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat
     4381 cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc
     4441 cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc
     4501 gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt
     4561 tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac
     4621 cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg
     4681 ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca
     4741 gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc
     4801 gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa
     4861 accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa
     4921 ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac
     4981 tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta
     5041 aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt
     5101 taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata
     5161 gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc
     5221 agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac
     5281 cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag
     5341 tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac
     5401 gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc
     5461 agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg
     5521 gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc
     5581 atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct
     5641 gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc
     5701 tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc
     5761 atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc
     5821 agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc
     5881 gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca
     5941 cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt
     6001 tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt
     6061 ccgcgcacat ttccccgaaa agtgccacct gacgtcttgt ttagcttgcc tcgtccccgc
     6121 cgggtcaccc ggccagcgac atggaggccc agaataccct ccttgacagt cttgacgtgc
     6181 gcagctcagg ggcatgatgt gactgtcgcc cgtacattta gcccatacat ccccatgtat
     6241 aatcatttgc atccatacat tttgatggcc gcacggcgcg aagcaaaaat tacggctcct
     6301 cgctgcagac ctgcgagcag ggaaacgctc ccctcacaga cgcgttgaat tgtccccacg
     6361 ccgcgcccct gtagagaaat ataaaaggtt aggatttgcc actgaggttc ttctttcata
     6421 tacttccttt taaaatcttg ctaggataca gttctcacat cacatccgaa cataaacaac
     6481 catgggtacc actcttgacg acacggctta ccggtaccgc accagtgtcc cgggggacgc
     6541 cgaggccatc gaggcactgg atgggtcctt caccaccgac accgtcttcc gcgtcaccgc
     6601 caccggggac ggcttcaccc tgcgggaggt gccggtggac ccgcccctga ccaaggtgtt
     6661 ccccgacgac gaatcggacg acgaatcgga cgacggggag gacggcgacc cggactcccg
     6721 gacgttcgtc gcgtacgggg acgacggcga cctggcgggc ttcgtggtcg tctcgtactc
     6781 cggctggaac cgccggctga ccgtcgagga catcgaggtc gccccggagc accgggggca
     6841 cggggtcggg cgcgcgttga tggggctcgc gacggagttc gcccgcgagc ggggcgccgg
     6901 gcacctctgg ctggaggtca ccaacgtcaa cgcaccggcg atccacgcgt accggcggat
     6961 ggggttcacc ctctgcggcc tggacaccgc cctgtacgac ggcaccgcct cggacggcga
     7021 gcaggcgctc tacatgagca tgccctgccc ctaatcagta ctgacaataa aaagattctt
     7081 gttttcaaga acttgtcatt tgtatagttt ttttatattg tagttgttct attttaatca
     7141 aatgttagcg tgatttatat tttttttcgc ctcgacatca tctgcccaga tgcgaagtta
     7201 agtgcgcaga aagtaatatc atgcgtcaat cgtatgtgaa tgctggtcgc tatactgctg
     7261 tcgattcgat actaacgccg ccatccagat catgacatta acctataaaa ataggcgtat
     7321 cacgaggccc tttcgtctcg cgcgtttcgg tgatgacggt gaaaacctct gacacatgca
     7381 gctcccggag acggtcacag cttgtctgta agcgg
//
