LOCUS       Exported                9271 bp ds-DNA     circular SYN 31-JUL-2023
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9271)
  AUTHORS   Lewis Lab
  TITLE     Direct Submission
  JOURNAL   Exported Monday, Dec 16, 2024 from SnapGene 5.0.8
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..9271
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     promoter        26..130
                     /label=AmpR promoter
                     /note="pLannotate"
                     /note="/database=snapgene"
                     /note="/fragment=False"
                     /note="/identity=100.0"
                     /note="/match_length=100.0"
                     /note="/other=promoter"
     CDS             131..991
                     /codon_start=1
                     /label=AmpR
                     /note="pLannotate"
                     /note="/database=snapgene"
                     /note="/fragment=False"
                     /note="/identity=99.8"
                     /note="/match_length=100.0"
                     /note="/other=CDS"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      1162..1750
                     /label=ori
                     /note="pLannotate"
                     /note="/database=snapgene"
                     /note="/fragment=False"
                     /note="/identity=99.8"
                     /note="/match_length=100.0"
                     /note="/other=rep_origin"
     primer_bind     complement(1796..1826)
                     /label=Z3_CRISP_V_R
     primer_bind     1807..1833
                     /label=Z3_CRISP_Ins_F
     misc_feature    1821..3841
                     /label=ACT1pr-yZ3EV-3'UTR
                     /note="ACT1pr-yZ3EV-3'UTR"
                     /note="Yeast codon-optimized Z3EV transcription factor
                     under the control of the ACT1 promoter; has the A. gossypii
                     TEF3 3'UTR"
     promoter        3848..4546
                     /label=Z3EV promoter
                     /note="Z3pr"
                     /note="Z3 promoter (Z3EV binding sites have replaced the
                     Gal4 binding sites in this modified GAL1 promoter)"
     primer_bind     complement(4501..4556)
                     /label=Z3_CRISP_Ins_R
     primer_bind     4537..4565
                     /label=Z3_CRISP_V_F
     primer_bind     4554..4573
                     /label=CRISPEY Retron SEQ F primer
                     /note="For sequence verifying inserted gRNA/repair 
                     templates"
     misc_feature    4591..4688
                     /label=Retron Ec86
     misc_feature    4689..4694
                     /label=XhoI cloning site
     misc_feature    4695..4856
                     /label=sgRNA and HDV
                     /note="structural guide RNA sequence and HDV ribozyme"
     primer_bind     complement(4695..4734)
                     /label=CRISPEY-R primer
                     /note="For amplification of ultramer/gBlock containing 
                     gRNA/repair template for cloning into CRISPEY vectors"
     misc_feature    4857..5212
                     /label=tGAL7
                     /note="GAL7 terminator"
     primer_bind     complement(5226..5244)
                     /label=T7 promoter primer
     primer_bind     complement(5254..5270)
                     /label=M13 F primer
     misc_feature    complement(5393..6749)
                     /label=KANMX cassette
     terminator      complement(5393..5590)
                     /label=TEF terminator
                     /note="pLannotate"
                     /note="/database=snapgene"
                     /note="/fragment=False"
                     /note="/identity=100.0"
                     /note="/match_length=100.0"
                     /note="/other=terminator"
     CDS             complement(5596..6405)
                     /codon_start=1
                     /label=KanR
                     /note="pLannotate"
                     /note="/database=snapgene"
                     /note="/fragment=False"
                     /note="/identity=100.0"
                     /note="/match_length=100.0"
                     /note="/other=CDS"
                     /translation="MGKEKTHVSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     promoter        complement(6406..6749)
                     /label=TEF promoter
                     /note="pLannotate"
                     /note="/database=snapgene"
                     /note="/fragment=False"
                     /note="/identity=99.7"
                     /note="/match_length=100.0"
                     /note="/other=promoter"
     rep_origin      6891..7319
                     /label=f1 ori
                     /note="pLannotate"
                     /note="/database=snapgene"
                     /note="/fragment=False"
                     /note="/identity=100.0"
                     /note="/match_length=100.0"
                     /note="/other=rep_origin"
     CDS             complement(7477..8280)
                     /codon_start=1
                     /label=URA3
                     /note="pLannotate"
                     /note="/database=snapgene"
                     /note="/fragment=False"
                     /note="/identity=99.8"
                     /note="/match_length=100.0"
                     /note="/other=CDS"
                     /translation="MSKATYKERAATHPSPVAAKLFNIMHEKQTNLCASLDVRTTKELL
                     ELVEALGPKICLLKTHVDILTDFSMEGTVKPLKALSAKYNFLLFEDRKFADIGNTVKLQ
                     YSAGVYRIAEWADITNAHGVVGPGIVSGLKQAAEEVTKEPRGLLMLAELSCKGSLSTGE
                     YTKGTVDIAKSDKDFVIGFIAQRDMGGRDEGYDWLIMTPGVGLDDKGDALGQQYRTVDD
                     VVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGQQN"
     promoter        complement(8281..8496)
                     /label=URA3 promoter
                     /note="pLannotate"
                     /note="/database=snapgene"
                     /note="/fragment=False"
                     /note="/identity=100.0"
                     /note="/match_length=100.0"
                     /note="/other=promoter"
     misc_feature    complement(8758..9260)
                     /label=CEN/ARS
                     /note="pLannotate"
                     /note="/database=snapgene"
                     /note="/fragment=False"
                     /note="/identity=99.8"
                     /note="/match_length=100.0"
                     /note="/other=misc_feature"
ORIGIN
        1 aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca
       61 ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa
      121 aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt ttgcggcatt
      181 ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg ctgaagatca
      241 gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga tccttgagag
      301 ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc tatgtggcgc
      361 ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac actattctca
      421 gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg gcatgacagt
      481 aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca acttacttct
      541 gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg gggatcatgt
      601 aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg acgagcgtga
      661 caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg gcgaactact
      721 tactctagct tcccggcaac aattaataga ctggatggag gcggataaag ttgcaggacc
      781 acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg gagccggtga
      841 gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct cccgtatcgt
      901 agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac agatcgctga
      961 gataggtgcc tcactgatta agcattggta actgtcagac caagtttact catatatact
     1021 ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga tcctttttga
     1081 taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt
     1141 agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca
     1201 aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct
     1261 ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc ttctagtgta
     1321 gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct
     1381 aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc
     1441 aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca
     1501 gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga
     1561 aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg
     1621 aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt
     1681 cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag
     1741 cctatggaaa aaggccagca acgcggcctt tttacggttc ctggcctttt gctggccttt
     1801 tgctggcctt ttgctcgctc gcgtcgatct cccctcaagt ggcaaactgc gactgatatg
     1861 aaagtaaagt atcggacctt ctattacgga atgtatacaa acgctacagg atacagtgat
     1921 attaaataga aaacaagtct acaattaaat ggcagaaaga agaaccagct tactactaac
     1981 caccatactc atcaataccc agagcatcgg tgaacatctg ctcaaattcg aagtcagcca
     2041 tatccagcgc accgtaaggt gctgaatcat gtggagtgaa gccaggccct ggagagtccc
     2101 cgtcacccaa catatctaaa tcaaaatcgt ctaaagcgtc agcgtgggcc atggcaacgt
     2161 cttcaccgtc caaatgcagt tcgctagatg tagaccctgc tgtcgctagg tgggattggt
     2221 cggtttcttc aacggaggct ccgcctctag aagttggtgc atgcaacctg tgagcgtcca
     2281 gcatttctag taataaatca tacagaggaa cgacgttttt acacttcata gaatacaggt
     2341 gctccatacc cttgttagac atatgtctaa tgtgtgacaa aattagtagt aactgagcta
     2401 gcctctgatg ctgttgctgc agagtcaatc ctgctttcgc cataaggtga atcaatgtat
     2461 cagtgatctt gtccaacact ctatgtatat ggtctttctc ttccaggctt ttcagagttg
     2521 agcttaaaaa tgtatagaca ccgctattta acaaaataat tgatttcagg caaacgaatt
     2581 cttctccttg caaattcatc attctgaacc tggagctagt ggcaagaagc atatcaaaaa
     2641 tctcaaccat accttctacg catttacctt ggttcctgtc caataataaa ttaggtgcaa
     2701 ataataattt tacagggtgc tccatacttc tccacactaa tccgatcatc aatatttcta
     2761 accacgcgca ttcaagaaga tgtacctggt catgtaatgt caagtcaaca aagccgggaa
     2821 ctcttttggc ccaattaatc atatgaacta attccctgtc tgctagattt gtcaaaagac
     2881 ccatcatgga agcctcggaa aaaggacgag ttgggtcgta ttcactgtac aatataggtg
     2941 gttcagcatc aagtagtgcg gaaaccattt gatcagcggt caaagacagt gcaagagaat
     3001 tcttcttaga tcttttgatc attaatgggc ttggccacaa gttggctgcc ctcatgtctc
     3061 cagcgcttgg atcttctagg gttgaggctg ctgccggagt gcccccacct cctgtatgga
     3121 tctttgtatg tctctttctt tcatcagatc tggcgaactt tcttccacaa atgtcacacg
     3181 cgaacggctt ctcgccggtg tgggttctaa tgtgcgtagt caaatggtca cttctgctga
     3241 agtttctcat acagattcta cattggaaag gcttttggcc ggtgtgaata cgaatatgcc
     3301 tagtcagctc gtcagatctg ctgaatctcc tatcacaact ttctacaggg caagcgtagg
     3361 gtctcgttcc cattgttaat tcagtaaatt ttcgatcttg ggaagaaaaa agcagtaagc
     3421 gtgaaaaatc taaaagctga tgtagtagaa gatcctattc tttaacaaag attgaccttt
     3481 tctttttctt cttggtttga gtagaaaggg gaaggaagaa tacaagagag aggaaaaaaa
     3541 ggaagataaa aagagagcgt gatataaatg aatatatatt aaacaagaga gattgggaag
     3601 gaaaggatca aacaaaccca aaaatatttc aaaaaggaga gagagaggcg agtttggttt
     3661 caaaacggtt tatttattta tgcaagagga cgtggaagaa aaagaagaag gaagaaaaaa
     3721 atttgaaaga aaaaaacgcg tggcgggtaa agaagaaaat ggaaaataga ggccgggtga
     3781 cagagaaata ttgagggtta attggaaaat atgttagggt gaggcatatg tttttaaagg
     3841 gactagttta tattgaattt tcaaaaattc ttactttttt tttggatgga cgcaaagaag
     3901 tttaataatc atattacatg gcattaccac catatacata tccatataca tatccatatc
     3961 taatcttact tatatgttgt ggaaatgtaa agagccccat tatcttagcc taaaaaaacc
     4021 ttctctttgg aactttcagt aatacgctta actgctcatt gctatattga agtgcggccg
     4081 cgtgggcgtg cgtgggcggg cgtgggcgtg cgtgggcggg cgtgggcgtg cgtgggcgtc
     4141 tagaccgtgc gtcctcgtct tcaccggtcg cgttcctgaa acgcagatgt gcctcgcgcc
     4201 gcactgctcc gaacaataaa gattctacaa tactagcttt tatggttatg aagaggaaaa
     4261 attggcagta acctggcccc acaaaccttc aaattaacga atcaaattaa caaccatagg
     4321 atgataatgc gattagtttt ttagccttat ttctggggta attaatcagc gaagcgatga
     4381 tttttgatct attaacagat atataaatgg aaaagctgca taaccacttt aactaatact
     4441 ttcaacattt tcagtttgta ttacttctta ttcaaatgtc ataaaagtat caacaaaaaa
     4501 ttgttaatat acctctatac tttaacgtca aggagaaaaa actataaggg tgcgcatctg
     4561 atgagtccgt gaggacgaaa cgagctagct cgtcatgcgc acccttagcg agaggtttat
     4621 cattaaggtc aacctctgga tgttgtttcg gcatcctgca ttgaatctga gttactgtct
     4681 gttttcctct cgaggtttca gagctatgct ggaaacagca tagcaagttg aaataaggct
     4741 agtccgttat caacttgaaa aagtggcacc gagtcggtgc tttttgatgg ccggcatggt
     4801 cccagcctcc tcgctggcgc cggctgggca acaccttcgg gtggcgaatg ggacttaaag
     4861 aaagtggaat attcattcat atcatatttt ttctattaac tgcctggttt cttttaaatt
     4921 ttttattggt tgtcgacttg aacggagtga caatatatat atatatattt aataatgaca
     4981 tcattatctg taaatctgat tcttaatgct attctagtta tgtaagagtg gtcctttcca
     5041 taaaaaaaaa aaaaagaaaa aagaatttta ggaatacaat gcagcttgta agtaaaatct
     5101 ggaatattca tatcgccaca acttcttatg cttataaaag cactaatgcc tgaatttatg
     5161 ttgaaaatat gtgtcacaaa taaagaaact gtgacatctg acacatttcc acgtacccaa
     5221 ttcgccctat agtgagtcgt attacgcgcg ctcactggcc gtcgttttac aacgtcgtga
     5281 ctgggaaaac cctggcgtta cccaacttaa tcgccttgca gcacatcccc ctttcgccag
     5341 aattcgagct cgttttcgac actggatggc ggcgttagta tcgaatcgac agcagtatag
     5401 cgaccagcat tcacatacga ttgacgcatg atattacttt ctgcgcactt aacttcgcat
     5461 ctgggcagat gatgtcgagg cgaaaaaaaa tataaatcac gctaacattt gattaaaata
     5521 gaacaactac aatataaaaa aactatacaa atgacaagtt cttgaaaaca agaatctttt
     5581 tattgtcagt actgattaga aaaactcatc gagcatcaaa tgaaactgca atttattcat
     5641 atcaggatta tcaataccat atttttgaaa aagccgtttc tgtaatgaag gagaaaactc
     5701 accgaggcag ttccatagga tggcaagatc ctggtatcgg tctgcgattc cgactcgtcc
     5761 aacatcaata caacctatta atttcccctc gtcaaaaata aggttatcaa gtgagaaatc
     5821 accatgagtg acgactgaat ccggtgagaa tggcaaaagc ttatgcattt ctttccagac
     5881 ttgttcaaca ggccagccat tacgctcgtc atcaaaatca ctcgcatcaa ccaaaccgtt
     5941 attcattcgt gattgcgcct gagcgagacg aaatacgcga tcgctgttaa aaggacaatt
     6001 acaaacagga atcgaatgca accggcgcag gaacactgcc agcgcatcaa caatattttc
     6061 acctgaatca ggatattctt ctaatacctg gaatgctgtt ttgccgggga tcgcagtggt
     6121 gagtaaccat gcatcatcag gagtacggat aaaatgcttg atggtcggaa gaggcataaa
     6181 ttccgtcagc cagtttagtc tgaccatctc atctgtaaca tcattggcaa cgctaccttt
     6241 gccatgtttc agaaacaact ctggcgcatc gggcttccca tacaatcgat agattgtcgc
     6301 acctgattgc ccgacattat cgcgagccca tttataccca tataaatcag catccatgtt
     6361 ggaatttaat cgcggcctcg aaacgtgagt cttttcctta cccatggttg tttatgttcg
     6421 gatgtgatgt gagaactgta tcctagcaag attttaaaag gaagtatatg aaagaagaac
     6481 ctcagtggca aatcctaacc ttttatattt ctctacaggg gcgcggcgtg gggacaattc
     6541 aacgcgtctg tgaggggagc gtttccctgc tcgcaggtcc gcagcgagga gccgtaattt
     6601 ttgcttcgcg ccgtgcggcc atcaaaatgt atggatgcaa atgattatac atggggatgt
     6661 atgggctaaa tgtacgggcg acagtcacat catgcccctg agctgcgcac gtcaagactg
     6721 tcaaggaggg tattctgggc ctccatgtcg ctggccgggt gacccggcgg ggacgaggca
     6781 agctaaacag atctggcgcg ccttaattaa cccggggatc ctggcgtaat agcgaagagg
     6841 cccgcaccga tcgcccttcc caacagttgc gcagcctgaa tggcgaatgg acgcgccctg
     6901 tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc
     6961 cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg
     7021 ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg
     7081 gcacctcgac cccaaaaaac ttgattaggg tgatggttca cgtagtgggc catcgccctg
     7141 atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt
     7201 ccaaactgga acaacactca accctatctc ggtctattct tttgatttat aagggatttt
     7261 gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta acgcgaattt
     7321 taacaaaata ttaacgctta caatttcctg atgcggtatt ttctccttac gcatctgtgc
     7381 ggtatttcac accgcatagg gtaataactg atataattaa attgaagctc taatttgtga
     7441 gtttagtata catgcattta cttataatac agttttttag ttttgctggc cgcatcttct
     7501 caaatatgct tcccagcctg cttttctgta acgttcaccc tctaccttag catcccttcc
     7561 ctttgcaaat agtcctcttc caacaataat aatgtcagat cctgtagaga ccacatcatc
     7621 cacggttcta tactgttgac ccaatgcgtc tcccttgtca tctaaaccca caccgggtgt
     7681 cataatcaac caatcgtaac cttcatctct tccacccatg tctctttgag caataaagcc
     7741 gataacaaaa tctttgtcgc tcttcgcaat gtcaacagta cccttagtat attctccagt
     7801 agatagggag cccttgcatg acaattctgc taacatcaaa aggcctctag gttcctttgt
     7861 tacttcttct gccgcctgct tcaaaccgct aacaatacct gggcccacca caccgtgtgc
     7921 attcgtaatg tctgcccatt ctgctattct gtatacaccc gcagagtact gcaatttgac
     7981 tgtattacca atgtcagcaa attttctgtc ttcgaagagt aaaaaattgt acttggcgga
     8041 taatgccttt agcggcttaa ctgtgccctc catggaaaaa tcagtcaaga tatccacatg
     8101 tgtttttagt aaacaaattt tgggacctaa tgcttcaact aactccagta attccttggt
     8161 ggtacgaaca tccaatgaag cacacaagtt tgtttgcttt tcgtgcatga tattaaatag
     8221 cttggcagca acaggactag gatgagtagc agcacgttcc ttatatgtag ctttcgacat
     8281 gatttatctt cgtttcctgc aggtttttgt tctgtgcagt tgggttaaga atactgggca
     8341 atttcatgtt tcttcaacac tacatatgcg tatatatacc aatctaagtc tgtgctcctt
     8401 ccttcgttct tccttctgtt cggagattac cgaatcaaaa aaatttcaag gaaaccgaaa
     8461 tcaaaaaaaa gaataaaaaa aaaatgatga attgaaaagg tggtatggtg cactctcagt
     8521 acaatctgct ctgatgccgc atagttaagc cagccccgac acccgccaac acccgctgac
     8581 gcgccctgac gggcttgtct gctcccggca tccgcttaca gacaagctgt gaccgtctcc
     8641 gggagctgca tgtgtcagag gttttcaccg tcatcaccga aacgcgcgag acgaaagggc
     8701 ctcgtgatac gcctattttt ataggttaat gtcatgataa taatggtttc ttagacggat
     8761 cgcttgcctg taacttacac gcgcctcgta tcttttaatg atggaataat ttgggaattt
     8821 actctgtgtt tatttatttt tatgttttgt atttggattt tagaaagtaa ataaagaagg
     8881 tagaagagtt acggaatgaa gaaaaaaaat aaacaaaggt ttaaaaaatt tcaacaaaaa
     8941 gcgtacttta catatatatt tattagacaa gaaaagcaga ttaaatagat atacattcga
     9001 ttaacgataa gtaaaatgta aaatcacagg attttcgtgt gtggtcttct acacagacaa
     9061 gatgaaacaa ttcggcatta atacctgaga gcaggaagag caagataaaa ggtagtattt
     9121 gttggcgatc cccctagagt cttttacatc ttcggaaaac aaaaactatt ttttctttaa
     9181 tttctttttt tactttctat ttttaattta tatatttata ttaaaaaatt taaattataa
     9241 ttatttttat agcacgtgat gaaaaggacc c
//
