LOCUS Exported 6585 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION EasyCloneYALI system-based yeast integrative vector carrying loxP-flanked nourseothricin-resistance marker, integration into Yarrowia lipolytica chromosomal location IntB, USER site for cloning, amp resistance. ACCESSION . VERSION . KEYWORDS pCfB4848 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6585) 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 6585) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6585 /organism="synthetic DNA construct" /mol_type="other DNA" protein_bind complement(1828..1861) /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." primer_bind complement(2979..2997) /label=CYC1 /note="CYC1 transcription termination signal, reverse primer" protein_bind complement(3196..3229) /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." protein_bind complement(3780..3813) /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(4020..4039) /label=pENTR-R /note="pENTR vectors, reverse primer" primer_bind complement(4045..4064) /label=Kan-R /note="Kanamycin resistance gene, reverse primer" promoter 4106..4197 /gene="bla" /label=AmpR promoter CDS 4198..5058 /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(4416..4435) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 5180..5768 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 5669..5688 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 5922..5939 /label=L4440 /note="L4440 vector, forward primer" primer_bind 6037..6054 /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind 6038..6054 /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" protein_bind 6524..6557 /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)" ORIGIN 1 gaatgcgtgc gatcccacag ttctcactca gatcatggag actctaacct tgagacatca 61 attatcagct ctcgaggata atgttagtgc agttccagga ctcattgtgc aactgtcacc 121 acggcatttt gggtctgttc ttttgaagta cagaaatatc ctcattgttg gtatactttg 181 ggactttttc ttgttacaga agaataaaaa aacctcgact gatgtactaa ttacatggtt 241 aacatcccca aggtcaaagt acagatattg taccgacttc tgaaatttgt gggatccaca 301 cacggacctc tgcgatgata caatatcatg gttccatggt ctcttgaatc acaccactca 361 ataataaaca ccagctcttt caatccaact tagctttctt ccacttgaac ataggccatc 421 cccccttctt tctattttac ataaatagca agatccttac ccctacatgt ctcataacac 481 aatctcaact tgacttccca taagaagttc actacgtgca acgcttacgc aactaacatg 541 aatgaatacg atatacatca aagactatga tacgcagtat tgcacactgt acgagtaagg 601 gcactagcca ctgcactcaa gtgaaaccgt tgcccgggta cgagtatgag tatgtacagt 661 atgtttagta ttgtacttgg acaatgcttg tatcgtacat tctcaagtgt caaacataaa 721 tatccgttgc tatatcctcg caccaccacg tagctcgcta tatccctgtg ttgaatccat 781 ccatcttgga ttgccaattg tgcacacaga accgggcact cacttcccca tccacacttc 841 aacggaatgc gtgcgatcgc gtgcattcct tctgttcgga atcaacctca aggttaacgg 901 ccacgatccc ctcgttgtta ctcttggtca gcccattgtc ggtaacgctg gctttgctaa 961 ctgggtcgat aaactcttct ttggccagga gaaccccgat gtctccaagg tgtccaaaga 1021 ccgaaagctc taccgaatca cccaccgagg agatatcgtc cctcaagtgc ccttctggga 1081 cggttaccag cactgctctg gtgaggtctt tattgactgg cccctgatcc accctcctct 1141 ctccaacgtt gtcatgtgcc agggccagag caataaacag tgctctgccg gtaacactct 1201 gctccagcag gtcaatgtga ttggaaacca tctgcagtac ttcgtcaccg agggtgtctg 1261 tggtatctaa gctatttatc actctttaca acttctacct caactatcta ctttaataaa 1321 tgaatatcgt ttattctcta tgattactgt atatgcgttc ctctaagaca aatcgaaacc 1381 agcatgcgat cgaatggcat acaaaagttt cttccgaagt tgatcaatgt cctgatagtc 1441 aggcagcttg agaagattga cacaggtgga ggccgtaggg aaccgatcaa cctgtctacc 1501 agcgttacga atggcaaatg acgggttcaa agccttgaat ccttgcaatg gtgccttgga 1561 tactgatgtc acaaacttaa gaagcagccg cttgtcctct tcctcgaaac tctcaaacac 1621 agtccagaag tcctttatag tttgatctgt atccagatag cctccgtaat tggtgtgtgt 1681 cttcaaatcc cagacgtcca cattggcatg tcctccactg ataagcattt gaagttcatc 1741 tgcgttgaac attgagaccc acgaagggtc aatgagctgg tatagaccgc ccaagaatgc 1801 atctgagaac agctgaagct tcgtacgata acttcgtata atgtatgcta tacgaagtta 1861 tagagaccgg gttggcggcg catttgtgtc ccaaaaaaca gccccaattg ccccaattga 1921 ccccaaattg acccagtagc gggcccaacc ccggcgagag cccccttctc cccacatatc 1981 aaacctcccc cggttcccac acttgccgtt aagggcgtag ggtactgcag tctggaatct 2041 acgcttgttc agactttgta ctagtttctt tgtctggcca tccgggtaac ccatgccgga 2101 cgcaaaatag actactgaaa atttttttgc tttgtggttg ggactttagc caagggtata 2161 aaagaccacc gtccccgaat tacctttcct cttcttttct ctctctcctt gtcaactcac 2221 acccgaaatc gttaagcatt tccttctgag tataagaatc attcaaaatg gtgagtttca 2281 gaggcagcag caattgccac gggctttgag cacacggccg ggtgtggtcc cattcccatc 2341 gacacaagac gccacgtcat ccgaccagca ctttttgcag tactaaccgc agggtactac 2401 tttggatgat actgcttaca gatacagaac ttctgttcca ggtgatgctg aagctattga 2461 agctttggat ggttctttca ctactgacac tgttttcaga gttactgcta ctggtgatgg 2521 tttcactttg agagaagttc cagttgatcc accattgact aaggtttttc cagatgatga 2581 atccgatgac gaatctgatg atggtgaaga tggtgatcca gattctagaa cttttgttgc 2641 ttatggtgat gacggtgact tggctggttt tgttgttgtt tcttattctg gttggaatag 2701 aagattgacc gtcgaagata ttgaagttgc tccagaacat agaggtcatg gtgttggtag 2761 agctttgatg ggtttggcta ctgaatttgc tagagaaaga ggtgctggtc atttgtggtt 2821 ggaagttact aatgttaacg ctccagctat tcatgcctat agaagaatgg gttttacctt 2881 gtgtggtttg gatactgcat tatacgatgg tactgcatca gatggtgaac aagccttgta 2941 tatgtctatg ccatgtccat gatcatgtaa ttagttatgt cacgcttaca ttcacgccct 3001 ccccccacat ccgctctaac cgaaaaggaa ggagttagac aacctgaagt ctaggtccct 3061 atttattttt ttatagttat gttagtatta agaacgttat ttatatttca aatttttctt 3121 ttttttctgt acagacgcgt gtacgcatgt aacattatac tgaaaacctt gcttgagaag 3181 gttttgggac gctcgataac ttcgtataat gtatgctata cgaagttatg tgatatcaga 3241 tccactagtg gcctacctgc tcctgcacct aagttcgtca aaatcggttt tactcagaga 3301 atcaacaacc ctaccaactg tacatactgc taaccctgat tctttgaata accccaataa 3361 ggctcctgct gacccttctg ccgttctagg aaaccagctt gttgagtacg ctgtgaacat 3421 cactcttggt atacccggtc agcccttctc agttcagatt gacacaggct cgtctgactt 3481 gtgggtgaag agtgacggct cctccggtgc attcaacaag aaggcttctt ctacttttca 3541 ggaggacgtt cccaacggct ttgcaattgc ccacggagac aaaacctctg ccattggaga 3601 ttgggtcaaa gataccatca atatcggtgg tgttagtatt gaccagtata gtattgacca 3661 gtatgagttc gccatggcta ctcagacaaa tactgacccg gtttttggta tcggctaccc 3721 gagcaacgag gcgtcttatg atcgcgtgca ttcgcggccg catttaaatc cccacttcag 3781 aagttcctat actttctaga gaataggaac ttcactatag agtcgaataa gggcgacacc 3841 ccataattag cccgggcgaa aggcccagtc tttcgactga gcctttcgtt ttatttgatg 3901 cctggcagtt ccctactctc gcatggggag tccccacact accatcggcg ctacggcgtt 3961 tcacttctga gttcggcatg gggtcaggtg ggaccaccgc gctactgccg ccaggcaaac 4021 aaggggtgtt atgagccata ttcaggtata aatgggctcg cgataatgtt cagaattggt 4081 taattggttg taacactgac ccctatttgt ttatttttct aaatacattc aaatatgtat 4141 ccgctcatga gacaataacc ctgataaatg cttcaataat attgaaaaag gaagaatatg 4201 agtattcaac atttccgtgt cgcccttatt cccttttttg cggcattttg ccttcctgtt 4261 tttgctcacc cagaaacgct ggtgaaagta aaagatgctg aagatcagtt gggtgcacga 4321 gtgggttaca tcgaactgga tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa 4381 gaacgttttc caatgatgag cacttttaaa gttctgctat gtggcgcggt attatcccgt 4441 attgacgccg ggcaagagca actcggtcgc cgcatacact attctcagaa tgacttggtt 4501 gagtactcac cagtcacaga aaagcatctt acggatggca tgacagtaag agaattatgc 4561 agtgctgcca taaccatgag tgataacact gcggccaact tacttctgac aacgatcgga 4621 ggaccgaagg agctaaccgc ttttttgcac aacatggggg atcatgtaac tcgccttgat 4681 cgttgggaac cggagctgaa tgaagccata ccaaacgacg agcgtgacac cacgatgcct 4741 gtagcgatgg caacaacgtt gcgcaaacta ttaactggcg aactacttac tctagcttcc 4801 cggcaacaat taatagactg gatggaggcg gataaagttg caggaccact tctgcgctcg 4861 gcccttccgg ctggctggtt tattgctgat aaatccggag ccggtgagcg tggttctcgc 4921 ggtatcatcg cagcgctggg gccagatggt aagccctccc gtatcgtagt tatctacacg 4981 acggggagtc aggcaactat ggatgaacga aatagacaga tcgctgagat aggtgcctca 5041 ctgattaagc attggtaagc agagcattac gctgacttga cgggacggcg caagctcatg 5101 accaaaatcc cttaacgtga gttacgcgcg cgtcgttcca ctgagcgtca gaccccgtag 5161 aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa 5221 caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt 5281 ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgttctt ctagtgtagc 5341 cgtagttagc ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa 5401 tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa 5461 gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc 5521 ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa 5581 gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa 5641 caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg 5701 ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc 5761 tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg 5821 ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg 5881 agtgagctga taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg 5941 aagcggaagg cgagagtagg gaactgccag gcatcaaact aagcagaagg cccctgacgg 6001 atggcctttt tgcgtttcta caaactcttt ctgtgttgta aaacgacggc cagtcttaag 6061 ctcgggcccc ctgggcggtt ctgataacga gtaatcgtta atccgcaaat aacgtaaaaa 6121 cccgcttcgg cgggtttttt tatgggggga gtttagggaa agagcatttg tcagaatatt 6181 taagggcgcc tgtcactttg cttgatatat gagaattatt taaccttata aatgagaaaa 6241 aagcaacgca ctttaaataa gatacgttgc tttttcgatt gatgaacacc tataattaaa 6301 ctattcatct attatttatg attttttgta tatacaatat ttctagtttg ttaaagagaa 6361 ttaagaaaat aaatctcgaa aataataaag ggaaaatcag tttttgatat caaaattata 6421 catgtcaacg ataatacaaa atataataca aactataaga tgttatcagt atttattatg 6481 catttagaat aaattttgtg tcgcccttat tcgactcact atagaagttc ctattctcta 6541 gaaagtatag gaacttcact tcattttatt taaatttgcg gccgc //