LOCUS       sequence-234453-        6708 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  ADE2-NatMx integrating (YIp) vector for stable genomic integration 
            in the BY4741 background..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6708)
  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 6708)
  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: YIp ADE2-NatMx
FEATURES             Location/Qualifiers
     source          1..6708
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(35..55)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(35..53)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(74..90)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(74..90)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(87..109)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    98..114
                     /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(122..128,129..146,147..152))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     primer_bind     complement(305..322)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(476..1064)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(556..575)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1235..2026,2027..2095))
                     /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     1858..1877
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2096..2200)
                     /gene="bla"
                     /label=AmpR promoter
     gene            2273..3395
                     /label=natMX6
                     /note="yeast selectable marker conferring nourseothricin 
                     resistance"
     promoter        2273..2616
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             2617..3192
                     /codon_start=1
                     /gene="Streptomyces noursei nat1"
                     /product="nourseothricin acetyltransferase"
                     /label=NrsR
                     /note="confers resistance to nourseothricin"
                     /translation="MGTTLDDTAYRYRTSVPGDAEAIEALDGSFTTDTVFRVTATGDGF
                     TLREVPVDPPLTKVFPDDESDDESDDGEDGDPDSRTFVAYGDDGDLAGFVVISYSAWNR
                     RLTVEDIEVAPEHRGHGVGRALMGLATEFAGERGAGHLWLEVTNVNAPAIHAYRRMGFT
                     LCGLDTALYDGTASDGERQALYMSMPCP"
     terminator      3198..3395
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     3448..3466
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(3504..3526)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     3626..3645
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        3674..4049
                     /gene="S. cerevisiae ADE2"
                     /label=ADE2 promoter
     CDS             4050..5765
                     /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(5970..6425)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6107..6128)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(join(6208..6708,1..78))
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSAQLTLTKGNKSWSSTAVAAALELVDPPGCRNSISSLSI
                     PSTSRGGPVPNSPYSESYYARSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEAR
                     TDRPSQQLRSLNGEWRDAPCSGALSAAGVVVTRSVTATLASALAPAPFAFFPSFLATFA
                     GFPRQALNRGLPLGFRFSALRHLDPKKLD"
     primer_bind     6319..6338
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6555..6577
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     6569..6586
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     6570..6586
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6596..6615
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        6596..6614
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_feature    join(6623..6708,1..22)
                     /label=MCS
                     /note="pBluescript multiple cloning site"
     primer_bind     6639..6655
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     6640..6656
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6690..6706)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6690..6706)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
ORIGIN
        1 ggccgccacc gcggtggagc tccagctttt gttcccttta gtgagggtta attgcgcgct
       61 tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac
      121 acaacatagg agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgaggtaac
      181 tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc
      241 tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg
      301 cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc
      361 actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt
      421 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc
      481 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa
      541 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc
      601 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg
      661 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc
      721 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
      781 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca
      841 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact
      901 acggctacac tagaaggaca gtatttggta tctgcgctct gctgaagcca gttaccttcg
      961 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt
     1021 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct
     1081 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga
     1141 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa
     1201 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac
     1261 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga
     1321 taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc
     1381 cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca
     1441 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta
     1501 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg
     1561 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc
     1621 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg
     1681 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt
     1741 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt
     1801 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata
     1861 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc
     1921 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac
     1981 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa
     2041 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct
     2101 tcctttttca atattattga agcatttatc agggttattg tctcatgagc ggatacatat
     2161 ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc
     2221 cacctgacgt cttgtttagc ttgcctcgtc cccgccgggt cacccggcca gcgacatgga
     2281 ggcccagaat accctccttg acagtcttga cgtgcgcagc tcaggggcat gatgtgactg
     2341 tcgcccgtac atttagccca tacatcccca tgtataatca tttgcatcca tacattttga
     2401 tggccgcacg gcgcgaagca aaaattacgg ctcctcgctg cagacctgcg agcagggaaa
     2461 cgctcccctc acagacgcgt tgaattgtcc ccacgccgcg cccctgtaga gaaatataaa
     2521 aggttaggat ttgccactga ggttcttctt tcatatactt ccttttaaaa tcttgctagg
     2581 atacagttct cacatcacat ccgaacataa acaaccatgg gtaccactct tgacgacacg
     2641 gcttaccggt accgcaccag tgtcccgggg gacgccgagg ccatcgaggc actggatggg
     2701 tccttcacca ccgacaccgt cttccgcgtc accgccaccg gggacggctt caccctgcgg
     2761 gaggtgccgg tggacccgcc cctgaccaag gtgttccccg acgacgaatc ggacgacgaa
     2821 tcggacgacg gggaggacgg cgacccggac tcccggacgt tcgtcgcgta cggggacgac
     2881 ggcgacctgg cgggcttcgt ggtcatctcg tactcggcgt ggaaccgccg gctgaccgtc
     2941 gaggacatcg aggtcgcccc ggagcaccgg gggcacgggg tcgggcgcgc gttgatgggg
     3001 ctcgcgacgg agttcgccgg cgagcggggc gccgggcacc tctggctgga ggtcaccaac
     3061 gtcaacgcac cggcgatcca cgcgtaccgg cggatggggt tcaccctctg cggcctggac
     3121 accgccctgt acgacggcac cgcctcggac ggcgagcggc aggcgctcta catgagcatg
     3181 ccctgcccct aatcagtact gacaataaaa agattcttgt tttcaagaac ttgtcatttg
     3241 tatagttttt ttatattgta gttgttctat tttaatcaaa tgttagcgtg atttatattt
     3301 tttttcgcct cgacatcatc tgcccagatg cgaagttaag tgcgcagaaa gtaatatcat
     3361 gcgtcaatcg tatgtgaatg ctggtcgcta tactgctgtc gattcgatac taacgccgcc
     3421 atccagatca tgacattaac ctataaaaat aggcgtatca cgaggccctt tcgtctcgcg
     3481 cgtttcggtg atgacggtga aaacctctga cacatgcagc tcccggagac ggtcacagct
     3541 tgtctgtaag cggatgccgg gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc
     3601 gggtgtcggg gctggcttaa ctatgcggca tcagagcaga ttgtactgag agtgcaccat
     3661 agatctgaat taattcttga ataatacata acttttctta aaagaatcaa agacagataa
     3721 aatttaagag atattaaata ttagtgagaa gccgagaatt ttgtaacacc aacataacac
     3781 tgacatcttt aacaactttt aattatgata catttcttac gtcatgattg attattacag
     3841 ctatgctgac aaatgactct tgttgcatgg ctacgaaccg ggtaatacta agtgattgac
     3901 tcttgctgac cttttattaa gaactaaatg gacaatatta tggagcattt catgtataaa
     3961 ttggtgcgta aaatcgttgg atctctcttc taagtacatc ctactataac aatcaagaaa
     4021 aacaagaaaa tcggacaaaa caatcaagta tggattctag aacagttggt atattaggag
     4081 ggggacaatt gggacgtatg attgttgagg cagcaaacag gctcaacatt aagacggtaa
     4141 tactagatgc tgaaaattct cctgccaaac aaataagcaa ctccaatgac cacgttaatg
     4201 gctccttttc caatcctctt gatatcgaaa aactagctga aaaatgtgat gtgctaacga
     4261 ttgagattga gcatgttgat gttcctacac taaagaatct tcaagtaaaa catcccaaat
     4321 taaaaattta cccttctcca gaaacaatca gattgataca agacaaatat attcaaaaag
     4381 agcatttaat caaaaatggt atagcagtta cccaaagtgt tcctgtggaa caagccagtg
     4441 agacgtccct attgaatgtt ggaagagatt tgggttttcc attcgtcttg aagtcgagga
     4501 ctttggcata cgatggaaga ggtaacttcg ttgtaaagaa taaggaaatg attccggaag
     4561 ctttggaagt actgaaggat cgtcctttgt acgccgaaaa atgggcacca tttactaaag
     4621 aattagcagt catgattgtg aggtctgtta acggtttagt gttttcttac ccaattgtag
     4681 agactatcca caaggacaat atttgtgact tatgttatgc gcctgctaga gttccggact
     4741 ccgttcaact taaggcgaag ttgttggcag aaaatgcaat caaatctttt cccggttgtg
     4801 gtatatttgg tgtggaaatg ttctatttag aaacagggga attgcttatt aacgaaattg
     4861 ccccaaggcc tcacaactct ggacattata ccattgatgc ttgcgtcact tctcaatttg
     4921 aagctcattt gagatcaata ttggatttgc caatgccaaa gaatttcaca tctttctcca
     4981 ccattacaac gaacgccatt atgctaaatg ttcttggaga caaacataca aaagataaag
     5041 agctagaaac ttgcgaaaga gcattggcga ctccaggttc ctcagtgtac ttatatggaa
     5101 aagagtctag acctaacaga aaagtaggtc acataaatat tattgcctcc agtatggcgg
     5161 aatgtgaaca aaggctgaac tacattacag gtagaactga tattccaatc aaaatctctg
     5221 tcgctcaaaa gttggacttg gaagcaatgg tcaaaccatt ggttggaatc atcatgggat
     5281 cagactctga cttgccggta atgtctgccg catgtgcggt tttaaaagat tttggcgttc
     5341 catttgaagt gacaatagtc tctgctcata gaactccaca taggatgtca gcatatgcta
     5401 tttccgcaag caagcgtgga attaaaacaa ttatcgctgg agctggtggg gctgctcact
     5461 tgccaggtat ggtggctgca atgacaccac ttcctgtcat cggtgtgccc gtaaaaggtt
     5521 cttgtctaga tggagtagat tctttacatt caattgtgca aatgcctaga ggtgttccag
     5581 tagctaccgt cgctattaat aatagtacga acgctgcgct gttggctgtc agactgcttg
     5641 gcgcttatga ttcaagttat acaacgaaaa tggaacagtt tttattaaag caagaagaag
     5701 aagttcttgt caaagcacaa aagttagaaa ctgtcggtta cgaagcttat ctagaaaaca
     5761 agtaatatat aagtttattg atatacttgt acagcaaata attataaaat gatataccta
     5821 ttttttaggc tttgttatga ttacatcaaa tgtggacttc atacatagaa atcaacgctt
     5881 acaggtgtcc ttttttaaga atttcataca taagatctat gcggtgtgaa ataccgcaca
     5941 gatgcgtaag gagaaaatac cgcatcagga aattgtaaac gttaatattt tgttaaaatt
     6001 cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat
     6061 cccttataaa tcaaaagaat agaccgagat agggttgagt gttgttccag tttggaacaa
     6121 gagtccacta ttaaagaacg tggactccaa cgtcaaaggg cgaaaaaccg tctatcaggg
     6181 cgatggccca ctacgtgaac catcacccta atcaagtttt ttggggtcga ggtgccgtaa
     6241 agcactaaat cggaacccta aagggagccc ccgatttaga gcttgacggg gaaagccggc
     6301 gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg ggcgctaggg cgctggcaag
     6361 tgtagcggtc acgctgcgcg taaccaccac acccgccgcg cttaatgcgc cgctacaggg
     6421 cgcgtcgcgc cattcgccat tcaggctgcg caactgttgg gaagggcgat cggtgcgggc
     6481 ctcttcgcta ttacgccagc tggcgaaagg gggatgtgct gcaaggcgat taagttgggt
     6541 aacgccaggg ttttcccagt cacgacgttg taaaacgacg gccagtgagc gcgcgtaata
     6601 cgactcacta tagggcgaat tgggtaccgg gccccccctc gaggtcgacg gtatcgataa
     6661 gcttgatatc gaattcctgc agcccggggg atccactagt tctagagc
//
