LOCUS       Exported                6767 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  N-terminal 9myc-AID* degron cassette with RFA1 promoter and KanR 
            marker for S. cerevisiae.
ACCESSION   .
VERSION     .
KEYWORDS    pKan-PRFA1-9myc-AID*(N)
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6767)
  AUTHORS   Morawska M, Ulrich HD
  TITLE     An expanded tool kit for the auxin-inducible degron system in 
            budding yeast.
  JOURNAL   Yeast. 2013 Sep;30(9):341-51. doi: 10.1002/yea.2967. Epub 2013 Jul 
            23.
  PUBMED    23836714
REFERENCE   2  (bases 1 to 6767)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6767
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     CDS             438..467
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             474..503
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             510..539
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             558..587
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             594..623
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             630..659
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             678..707
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             714..743
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             750..779
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     promoter        complement(1035..1053)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(1074..1090)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1074..1090)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1087..1109)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1098..1114
                     /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(1122..1152)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1167..1188
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1305..1322)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1476..2064)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1556..1575)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(2235..3095)
                     /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     2858..2877
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3096..3200)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3268..3286
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(3324..3346)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     3446..3465
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        3489..3704
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     CDS             3705..4508
                     /codon_start=1
                     /gene="S. cerevisiae URA3"
                     /product="orotidine-5'-phosphate decarboxylase, required 
                     for uracil biosynthesis"
                     /label=URA3
                     /note="yeast auxotrophic marker, counterselectable with 
                     5-fluoroorotic acid (5-FOA)"
                     /translation="MSKATYKERAATHPSPVAAKLFNIMHEKQTNLCASLDVRTTKELL
                     ELVEALGPKICLLKTHVDILTDFSMEGTVKPLKALSAKYNFLLFEDRKFADIGNTVKLQ
                     YSAGVYRIAEWADITNAHGVVGPGIVSGLKQAAEEVTKEPRGLLMLAELSCKGSLSTGE
                     YTKGTVDIAKSDKDFVIGFIAQRDMGGRDEGYDWLIMTPGVGLDDKGDALGQQYRTVDD
                     VVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGQQN"
     rep_origin      complement(4639..5094)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4776..4797)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4988..5007
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5220..5242
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     5234..5251
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     5235..5251
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5258..5277
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5258..5276
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     gene            5411..6767
                     /label=kanMX
                     /note="yeast selectable marker conferring kanamycin 
                     resistance (Wach et al., 1994)"
     promoter        5411..5754
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             5755..6564
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin"
                     /translation="MGKEKTHVSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(5822..5841)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     terminator      6570..6767
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
ORIGIN
        1 ctgtcgattc gatactaacg ccgccatcca gtgtcgaaaa cgaggcggcc gctctagaac
       61 tagtgctagc tcgccttttg ccgttctttt tgcatatatg ttaatagcta tgcgcgtacc
      121 accttcttca gccatatacg aaaggatcta tgctaattgg gtggatttct tatcgcatca
      181 ctttcttatt ctctaactct tcttcgctgc tcaataaaat atctctgctt ggaaaaatgt
      241 cacgcgtaaa aattagacga ggcgacaaaa gacgcgtgaa ctaacataaa ggtttacttg
      301 aacaaatgtt atcactttaa gttctacacg taaaacaaaa actattcact cttcttatta
      361 cggtaaaggc gaaaccagca agaagaccag attatactta caagacacgt gtcatgtccg
      421 gttctgctgc tagtggtgaa caaaagttga tttctgaaga agatttgaac ggtgaacaaa
      481 agctaatctc cgaggaagac ttgaacggtg aacaaaaatt aatctcagaa gaagacttga
      541 acggatcctc tagaggtgaa caaaagttga tttctgaaga agatttgaac ggtgaacaaa
      601 agctaatctc cgaggaagac ttgaacggtg aacaaaaatt aatctcagaa gaagacttga
      661 acggatcctc tagaggtgaa caaaagttga tttctgaaga agatttgaac ggtgaacaaa
      721 agctaatctc cgaggaagac ttgaacggtg aacaaaaatt aatctcagaa gaagacttga
      781 acggatccac tagagaattc atgcctaaag atccagccaa acctccggcc aaggcacaag
      841 ttgtgggatg gccaccggtg agatcatacc ggaagaacgt gatggtttcc tgccaaaaat
      901 caagcggtgg cccggaggcg gcggcgttcg tgaaggtatc aatgctcgag gaagaacttc
      961 aaattgaatc taaggacttg ggagctggtg caggcgctgg agcgggtgcc agagctcggt
     1021 accagctttt gttcccttta gtgagggtta atttcgagct tggcgtaatc atggtcatag
     1081 ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc
     1141 ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc
     1201 tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa
     1261 cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg
     1321 ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg
     1381 ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag
     1441 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
     1501 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
     1561 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
     1621 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
     1681 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     1741 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
     1801 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
     1861 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca
     1921 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
     1981 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
     2041 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
     2101 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc
     2161 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa
     2221 acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
     2281 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
     2341 ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat
     2401 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
     2461 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
     2521 aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt
     2581 ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg
     2641 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc
     2701 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc
     2761 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg
     2821 cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga
     2881 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta
     2941 ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct
     3001 tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag
     3061 ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca atattattga
     3121 agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat
     3181 aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgt ctaagaaacc
     3241 attattatca tgacattaac ctataaaaat aggcgtatca cgaggccctt tcgtctcgcg
     3301 cgtttcggtg atgacggtga aaacctctga cacatgcagc tcccggagac ggtcacagct
     3361 tgtctgtaag cggatgccgg gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc
     3421 gggtgtcggg gctggcttaa ctatgcggca tcagagcaga ttgtactgag agtgcaccat
     3481 accacctttt caattcatca tttttttttt attctttttt ttgatttcgg tttccttgaa
     3541 atttttttga ttcggtaatc tccgaacaga aggaagaacg aaggaaggag cacagactta
     3601 gattggtata tatacgcata tgtagtgttg aagaaacatg aaattgccca gtattcttaa
     3661 cccaactgca cagaacaaaa acctgcagga aacgaagata aatcatgtcg aaagctacat
     3721 ataaggaacg tgctgctact catcctagtc ctgttgctgc caagctattt aatatcatgc
     3781 acgaaaagca aacaaacttg tgtgcttcat tggatgttcg taccaccaag gaattactgg
     3841 agttagttga agcattaggt cccaaaattt gtttactaaa aacacatgtg gatatcttga
     3901 ctgatttttc catggagggc acagttaagc cgctaaaggc attatccgcc aagtacaatt
     3961 ttttactctt cgaagacaga aaatttgctg acattggtaa tacagtcaaa ttgcagtact
     4021 ctgcgggtgt atacagaata gcagaatggg cagacattac gaatgcacac ggtgtggtgg
     4081 gcccaggtat tgttagcggt ttgaagcagg cggcagaaga agtaacaaag gaacctagag
     4141 gccttttgat gttagcagaa ttgtcatgca agggctccct atctactgga gaatatacta
     4201 agggtactgt tgacattgcg aagagcgaca aagattttgt tatcggcttt attgctcaaa
     4261 gagacatggg tggaagagat gaaggttacg attggttgat tatgacaccc ggtgtgggtt
     4321 tagatgacaa gggagacgca ttgggtcaac agtatagaac cgtggatgat gtggtctcta
     4381 caggatctga cattattatt gttggaagag gactatttgc aaagggaagg gatgctaagg
     4441 tagagggtga acgttacaga aaagcaggct gggaagcata tttgagaaga tgcggccagc
     4501 aaaactaaaa aactgtatta taagtaaatg catgtatact aaactcacaa attagagctt
     4561 caatttaatt atatcagtta ttaccctatg cggtgtgaaa taccgcacag atgcgtaagg
     4621 agaaaatacc gcatcaggaa attgtaagcg ttaatatttt gttaaaattc gcgttaaatt
     4681 tttgttaaat cagctcattt tttaaccaat aggccgaaat cggcaaaatc ccttataaat
     4741 caaaagaata gaccgagata gggttgagtg ttgttccagt ttggaacaag agtccactat
     4801 taaagaacgt ggactccaac gtcaaagggc gaaaaaccgt ctatcagggc gatggcccac
     4861 tacgtgaacc atcaccctaa tcaagttttt tggggtcgag gtgccgtaaa gcactaaatc
     4921 ggaaccctaa agggagcccc cgatttagag cttgacgggg aaagccggcg aacgtggcga
     4981 gaaaggaagg gaagaaagcg aaaggagcgg gcgctagggc gctggcaagt gtagcggtca
     5041 cgctgcgcgt aaccaccaca cccgccgcgc ttaatgcgcc gctacagggc gcgtccattc
     5101 gccattcagg ctgcgcaact gttgggaagg gcgatcggtg cgggcctctt cgctattacg
     5161 ccagctggcg aaagggggat gtgctgcaag gcgattaagt tgggtaacgc cagggttttc
     5221 ccagtcacga cgttgtaaaa cgacggccag tgaattgtaa tacgactcac tatagggcga
     5281 attggagctc caccgcggtg gcggccgcca gctgaagctt cgtacgctgc aggtcgacgg
     5341 atccccgggt taattaaggc gcgccagatc tgtttagctt gcctcgtccc cgccgggtca
     5401 cccggccagc gacatggagg cccagaatac cctccttgac agtcttgacg tgcgcagctc
     5461 aggggcatga tgtgactgtc gcccgtacat ttagcccata catccccatg tataatcatt
     5521 tgcatccata cattttgatg gccgcacggc gcgaagcaaa aattacggct cctcgctgca
     5581 gacctgcgag cagggaaacg ctcccctcac agacgcgttg aattgtcccc acgccgcgcc
     5641 cctgtagaga aatataaaag gttaggattt gccactgagg ttcttctttc atatacttcc
     5701 ttttaaaatc ttgctaggat acagttctca catcacatcc gaacataaac aaccatgggt
     5761 aaggaaaaga ctcacgtttc gaggccgcga ttaaattcca acatggatgc tgatttatat
     5821 gggtataaat gggctcgcga taatgtcggg caatcaggtg cgacaatcta tcgattgtat
     5881 gggaagcccg atgcgccaga gttgtttctg aaacatggca aaggtagcgt tgccaatgat
     5941 gttacagatg agatggtcag actaaactgg ctgacggaat ttatgcctct tccgaccatc
     6001 aagcatttta tccgtactcc tgatgatgca tggttactca ccactgcgat ccccggcaaa
     6061 acagcattcc aggtattaga agaatatcct gattcaggtg aaaatattgt tgatgcgctg
     6121 gcagtgttcc tgcgccggtt gcattcgatt cctgtttgta attgtccttt taacagcgat
     6181 cgcgtatttc gtctcgctca ggcgcaatca cgaatgaata acggtttggt tgatgcgagt
     6241 gattttgatg acgagcgtaa tggctggcct gttgaacaag tctggaaaga aatgcataag
     6301 cttttgccat tctcaccgga ttcagtcgtc actcatggtg atttctcact tgataacctt
     6361 atttttgacg aggggaaatt aataggttgt attgatgttg gacgagtcgg aatcgcagac
     6421 cgataccagg atcttgccat cctatggaac tgcctcggtg agttttctcc ttcattacag
     6481 aaacggcttt ttcaaaaata tggtattgat aatcctgata tgaataaatt gcagtttcat
     6541 ttgatgctcg atgagttttt ctaatcagta ctgacaataa aaagattctt gttttcaaga
     6601 acttgtcatt tgtatagttt ttttatattg tagttgttct attttaatca aatgttagcg
     6661 tgatttatat tttttttcgc ctcgacatca tctgcccaga tgcgaagtta agtgcgcaga
     6721 aagtaatatc atgcgtcaat cgtatgtgaa tgctggtcgc tatactg
//
