LOCUS       Exported                5994 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  EasyCloneYALI system-based yeast episomal vector carrying a 
            nourseothricin-resistance marker for Yarrowia lipolytica, can be 
            used for gRNA expression, amp resistance.
ACCESSION   .
VERSION     .
KEYWORDS    pCfB3405
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5994)
  AUTHORS   Holkenbrink C, Dam MI, Kildegaard KR, Beder J, Dahlin J, Domenech 
            Belda D, Borodina I
  TITLE     EasyCloneYALI: CRISPR/Cas9-Based Synthetic Toolbox for Engineering 
            of the Yeast Yarrowia lipolytica.
  JOURNAL   Biotechnol J. 2018 Jan 29. doi: 10.1002/biot.201700543.
  PUBMED    29377615
REFERENCE   2  (bases 1 to 5994)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..5994
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     protein_bind    complement(19..52)
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     primer_bind     complement(259..278)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     primer_bind     complement(284..303)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        345..436
                     /gene="bla"
                     /label=AmpR promoter
     CDS             437..1297
                     /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     complement(655..674)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1419..2007
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1908..1927
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2161..2178
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     2276..2293
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2277..2293
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    2763..2796
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     primer_bind     4264..4282
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
ORIGIN
        1 atttaaatcc ccacttcaga agttcctata ctttctagag aataggaact tcactataga
       61 gtcgaataag ggcgacaccc cataattagc ccgggcgaaa ggcccagtct ttcgactgag
      121 cctttcgttt tatttgatgc ctggcagttc cctactctcg catggggagt ccccacacta
      181 ccatcggcgc tacggcgttt cacttctgag ttcggcatgg ggtcaggtgg gaccaccgcg
      241 ctactgccgc caggcaaaca aggggtgtta tgagccatat tcaggtataa atgggctcgc
      301 gataatgttc agaattggtt aattggttgt aacactgacc cctatttgtt tatttttcta
      361 aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata
      421 ttgaaaaagg aagaatatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc
      481 ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga
      541 agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct
      601 tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg
      661 tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta
      721 ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat
      781 gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt
      841 acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga
      901 tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga
      961 gcgtgacacc acgatgcctg tagcgatggc aacaacgttg cgcaaactat taactggcga
     1021 actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc
     1081 aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata aatccggagc
     1141 cggtgagcgt ggttctcgcg gtatcatcgc agcgctgggg ccagatggta agccctcccg
     1201 tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat
     1261 cgctgagata ggtgcctcac tgattaagca ttggtaagca gagcattacg ctgacttgac
     1321 gggacggcgc aagctcatga ccaaaatccc ttaacgtgag ttacgcgcgc gtcgttccac
     1381 tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc
     1441 gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat
     1501 caagagctac caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat
     1561 actgttcttc tagtgtagcc gtagttagcc caccacttca agaactctgt agcaccgcct
     1621 acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt
     1681 cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg
     1741 gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta
     1801 cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg
     1861 gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg
     1921 tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc
     1981 tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg
     2041 gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat
     2101 aaccgtatta ccgcctttga gtgagctgat accgctcgcc gcagccgaac gaccgagcgc
     2161 agcgagtcag tgagcgagga agcggaaggc gagagtaggg aactgccagg catcaaacta
     2221 agcagaaggc ccctgacgga tggccttttt gcgtttctac aaactctttc tgtgttgtaa
     2281 aacgacggcc agtcttaagc tcgggccccc tgggcggttc tgataacgag taatcgttaa
     2341 tccgcaaata acgtaaaaac ccgcttcggc gggttttttt atggggggag tttagggaaa
     2401 gagcatttgt cagaatattt aagggcgcct gtcactttgc ttgatatatg agaattattt
     2461 aaccttataa atgagaaaaa agcaacgcac tttaaataag atacgttgct ttttcgattg
     2521 atgaacacct ataattaaac tattcatcta ttatttatga ttttttgtat atacaatatt
     2581 tctagtttgt taaagagaat taagaaaata aatctcgaaa ataataaagg gaaaatcagt
     2641 ttttgatatc aaaattatac atgtcaacga taatacaaaa tataatacaa actataagat
     2701 gttatcagta tttattatgc atttagaata aattttgtgt cgcccttatt cgactcacta
     2761 tagaagttcc tattctctag aaagtatagg aacttctgaa gtggggattt aaatgcggcc
     2821 gcgaatgcac gcgatcgcac gcattccgtt gcatcaaatt tagggatgcc atcaactttc
     2881 agttcataat taatatctta ccaaattagg taatctgcaa aagttcagac tgtgaaatgt
     2941 aacattttat atatcaagct ctatttaatg cctcacagta gttaacataa agagatacag
     3001 aattgtcgtg tcagtgtata ctatccatgt gtatactctg gatatccatt tgtattccat
     3061 tatctacgaa ttggcggggt tctgctcgag ctgggtggcg aggttgttaa gtcggtccat
     3121 ttttgtgtgt cttggttgga tgaggtccaa agtgggggga gtctaccagg ctttatatac
     3181 acctggtagt tgcaactttc ggtatgatcc ccacgaaaga taaagttcag catgtggaga
     3241 gccgaggtag tatgcgtgtg acagagaccc aaactggggg tcaccgcgtg cacactggcg
     3301 acgccgctct ggaaggtcac atgaccgcag atactccgtt ctcccctcac gtcgattcgg
     3361 ctcggggaca cggtcagcgc cattgccacg tgtccacggt tctctccact ctgtagaaag
     3421 tcacttccac caagttccat gaagcttatt aggtttttgg agaaatccgg ggcggcattt
     3481 ctggggtaac ttgggcaagc cagtagtgga tcaattggga cccgaattgg gtggaaaaag
     3541 gtgataacga gccctttggg atccccacaa ttgatctaag acctctgctt gccccgatct
     3601 tggccccctc agccccaaca ctgttgctct tcaaactccc ccatacctcg gttcagatgc
     3661 agccttcaaa actttaacgt acctccgagc ctgtgaaaac gtggtcatga aaaaacgaga
     3721 gatgtaagtc ccgtcacata ccgctgcgag accaccagct gtacatacac ttggcgttcc
     3781 agcatgcacg gtcgctcaac aggcgctccc ccattggcgg aaagacaagc ccccactaga
     3841 ttcctgttca agggcaaagt ctagaagccc gcccccccca gcgcccgttt ccttgttagt
     3901 gcggcctgca catgtcacct gtctggcagg tatcgtcgct actttgccag tcgtatcgtc
     3961 gtcaacggat tgcgctgccc attcacgtgt aaactctcga cggatatccc ggagtttccc
     4021 gccgatcctc ccgcgagcca ctgccacgtc gcctccaagc ctgctcgagc gtcccaaaac
     4081 cttctcaagc aaggttttca gtataatgtt acatgcgtac acgcgtctgt acagaaaaaa
     4141 aagaaaaatt tgaaatataa ataacgttct taatactaac ataactataa aaaaataaat
     4201 agggacctag acttcaggtt gtctaactcc ttccttttcg gttagagcgg atgtgggggg
     4261 agggcgtgaa tgtaagcgtg acataactaa ttacatgatc atggacatgg catagacata
     4321 tacaaggctt gttcaccatc tgatgcagta ccatcgtata atgcagtatc caaaccacac
     4381 aaggtaaaac ccattcttct ataggcatga atagctggag cgttaacatt agtaacttcc
     4441 aaccacaaat gaccagcacc tctttctcta gcaaattcag tagccaaacc catcaaagct
     4501 ctaccaacac catgacctct atgttctgga gcaacttcaa tatcttcgac ggtcaatctt
     4561 ctattccaac cagaataaga aacaacaaca aaaccagcca agtcaccgtc atcaccataa
     4621 gcaacaaaag ttctagaatc tggatcacca tcttcaccat catcagattc gtcatcggat
     4681 tcatcatctg gaaaaacctt agtcaatggt ggatcaactg gaacttctct caaagtgaaa
     4741 ccatcaccag tagcagtaac tctgaaaaca gtgtcagtag tgaaagaacc atccaaagct
     4801 tcaatagctt cagcatcacc tggaacagaa gttctgtatc tgtaagcagt atcatccaaa
     4861 gtagtaccca ttgctgtaga tatgtcttgt gtgtaagggg gttggggtgg ttgtttgtgt
     4921 tcttgacttt tgtgttagca agggaagacg ggcaaaaaag tgagtgtggt tgggagggag
     4981 agacgagcct tatatataat gcttgtttgt gtttgtgcaa gtggacgccg aaacgggcag
     5041 gagccaaact aaacaaggca gacaatgcga gcttaattgg attgcctgat gggcaggggt
     5101 tagggctcga tcaatggggg tgcgaagtga caaaattggg aattaggttc gcaagcaagg
     5161 ctgacaagac tttggcccaa acatttgtac gcggtggaca acaggagcca cccatcgtct
     5221 gtcacgggct agccggtcgt gcgtcctgtc aggctccacc taggctccat gccactccat
     5281 acaatcccac tagtgtaccg ctaggccgct tttagctccc atctaagacc cccccaaaac
     5341 ctccactgta cagtgcactg tactgtgtgg cgatcaaggg caagggaaaa aaggcgcaaa
     5401 catgcatgca tggaatgacg taggtaaggc gttactagac tgaaaagtgg cacatttcgg
     5461 cgtgccaaag ggtcctaggt gcgtttcgcg agctgggcgc caggccaagc cgctccaaaa
     5521 cgcctctccg actccctcca gcggcctcca tatccccatc cctctccaca gcaatgttgt
     5581 taagccttgc aaacgaaaaa atagaaaggc taataagctt ccaatattgt ggtgtacgct
     5641 gcataacgca acaatgagcg ccaaacaaca cacacacaca gcacacagca gcattaacca
     5701 cgatgaacag catgacatta caggtgggtg tgtaatcagg gccctgattg ctggtggtgg
     5761 gagcccccat catgggctct cgtccagccg cggaaaccgg aaacgctcga atctgccgcc
     5821 gctcatactc accgaaacgt gtggggctcg aaaaaacggg cgccaaactc cttatactac
     5881 tgtatattca agcaagtata tccgtgggtg cgggtgattt ggatctaagg ttcgtactca
     5941 acactcacga gcagcttgcc tatgttacat ccttttatca gacatagcgg ccgc
//
