LOCUS       sequence-206240-        6586 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  EasyCloneYALI system-based yeast integrative vector carrying 
            loxP-flanked nourseothricin-resistance marker, integration into 
            Yarrowia lipolytica chromosomal location IntA_1, USER site for 
            cloning, amp resistance.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6586)
  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 6586)
  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: "Biotechnol 
            J. 2018 Jan 29. doi: 10.1002/biot.201700543."
COMMENT     Sequence Label: pCfB4788
FEATURES             Location/Qualifiers
     source          1..6586
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    1819..1852
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     complement(2970..2988)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     protein_bind    3187..3220
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    complement(3781..3814)
                     /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(4021..4040)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     primer_bind     complement(4046..4065)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        4107..4198
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4199..4267,4268..5059)
                     /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(4417..4436)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5181..5769
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5670..5689
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5923..5940
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     6038..6055
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     6039..6055
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    6525..6558
                     /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 gatttattat cccaaaaata ccccactctc cattatttcc gactgtgcag
       61 attcccatat caacccaatc tccatcaaaa caccgcagcc aaactctggc aaaagtgcaa
      121 catgcgatac ggacacggtg ggcggtgggc atcgatcaac tggccgtagg tgcatctggg
      181 ctggaattac cccacagtgc agtgctaagc agtggtgatg gaggatattg ggggatggaa
      241 gatattgggg gatggaggga gaggactaga cctctcagta acagtgcaag cagtacatgc
      301 aatacaagta acacccacca acacacggac cagacgagac caaccaactg cacagcccta
      361 ccaggaaacg gcagagcttg tggacacaac agactgcgac aagggacccg ctttgccagt
      421 gtgatgcagg aaccatactc ttttcctttt ttcggacagc cggcacacga gcttgacgat
      481 caaaccatat cggccgagtg cgacacgtgc aacgcttacg caactaacat gaatgaatac
      541 gatatacatc aaagactatg atacgcagta ttgcacactg tacgagtaag ggcactagcc
      601 actgcactca agtgaaaccg ttgcccgggt acgagtatga gtatgtacag tatgtttagt
      661 attgtacttg gacaatgctt gtatcgtaca ttctcaagtg tcaaacataa atatccgttg
      721 ctatatcctc gcaccaccac gtagctcgct atatccctgt gttgaatcca tccatcttgg
      781 attgccaatt gtgcacacag aaccgggcac tcacttcccc atccacactt caacggaatg
      841 cgtgcgatcg cgtgcattcc ttctgttcgg aatcaacctc aaggttaacg gccacgatcc
      901 cctcgttgtt actcttggtc agcccattgt cggtaacgct ggctttgcta actgggtcga
      961 taaactcttc tttggccagg agaaccccga tgtctccaag gtgtccaaag accgaaagct
     1021 ctaccgaatc acccaccgag gagatatcgt ccctcaagtg cccttctggg acggttacca
     1081 gcactgctct ggtgaggtct ttattgactg gcccctgatc caccctcctc tctccaacgt
     1141 tgtcatgtgc cagggccaga gcaataaaca gtgctctgcc ggtaacactc tgctccagca
     1201 ggtcaatgtg attggaaacc atctgcagta cttcgtcacc gagggtgtct gtggtatcta
     1261 agctatttat cactctttac aacttctacc tcaactatct actttaataa atgaatatcg
     1321 tttattctct atgattactg tatatgcgtt cctctaagac aaatcgaaac cagcatgcga
     1381 tcgaatggca tacaaaagtt tcttccgaag ttgatcaatg tcctgatagt caggcagctt
     1441 gagaagattg acacaggtgg aggccgtagg gaaccgatca acctgtctac cagcgttacg
     1501 aatggcaaat gacgggttca aagccttgaa tccttgcaat ggtgccttgg atactgatgt
     1561 cacaaactta agaagcagcc gcttgtcctc ttcctcgaaa ctctcaaaca cagtccagaa
     1621 gtcctttata gtttgatctg tatccagata gcctccgtaa ttggtgtgtg tcttcaaatc
     1681 ccagacgtcc acattggcat gtcctccact gataagcatt tgaagttcat ctgcgttgaa
     1741 cattgagacc cacgaagggt caatgagctg gtatagaccg cccaagaatg catctgagaa
     1801 cagctgaagc ttcgtacgat aacttcgtat aatgtatgct atacgaagtt atagagaccg
     1861 ggttggcggc gcatttgtgt cccaaaaaac agccccaatt gccccaattg accccaaatt
     1921 gacccagtag cgggcccaac cccggcgaga gcccccttct ccccacatat caaacctccc
     1981 ccggttccca cacttgccgt taagggcgta gggtactgca gtctggaatc tacgcttgtt
     2041 cagactttgt actagtttct ttgtctggcc atccgggtaa cccatgccgg acgcaaaata
     2101 gactactgaa aatttttttg ctttgtggtt gggactttag ccaagggtat aaaagaccac
     2161 cgtccccgaa ttacctttcc tcttcttttc tctctctcct tgtcaactca cacccgaaat
     2221 cgttaagcat ttccttctga gtataagaat cattcaaaat ggtgagtttc agaggcagca
     2281 gcaattgcca cgggctttga gcacacggcc gggtgtggtc ccattcccat cgacacaaga
     2341 cgccacgtca tccgaccagc actttttgca gtactaaccg cagggtacta ctttggatga
     2401 tactgcttac agatacagaa cttctgttcc aggtgatgct gaagctattg aagctttgga
     2461 tggttctttc actactgaca ctgttttcag agttactgct actggtgatg gtttcacttt
     2521 gagagaagtt ccagttgatc caccattgac taaggttttt ccagatgatg aatccgatga
     2581 cgaatctgat gatggtgaag atggtgatcc agattctaga acttttgttg cttatggtga
     2641 tgacggtgac ttggctggtt ttgttgttgt ttcttattct ggttggaata gaagattgac
     2701 cgtcgaagat attgaagttg ctccagaaca tagaggtcat ggtgttggta gagctttgat
     2761 gggtttggct actgaatttg ctagagaaag aggtgctggt catttgtggt tggaagttac
     2821 taatgttaac gctccagcta ttcatgccta tagaagaatg ggttttacct tgtgtggttt
     2881 ggatactgca ttatacgatg gtactgcatc agatggtgaa caagccttgt atatgtctat
     2941 gccatgtcca tgatcatgta attagttatg tcacgcttac attcacgccc tccccccaca
     3001 tccgctctaa ccgaaaagga aggagttaga caacctgaag tctaggtccc tatttatttt
     3061 tttatagtta tgttagtatt aagaacgtta tttatatttc aaatttttct tttttttctg
     3121 tacagacgcg tgtacgcatg taacattata ctgaaaacct tgcttgagaa ggttttggga
     3181 cgctcgataa cttcgtataa tgtatgctat acgaagttat gtgatatcag atccactagt
     3241 ggcctggggc tggcgtgtga aggagcatgt gcaaagggcc gcgcagactt gtggagtgct
     3301 gccgtacgag tatcggtgta cgacaaggtg gtttgtacaa ggtggtttgt accgtcggac
     3361 atgatcatct gtaccttggc gtactgtact tgtacaacca cttggtgatc agatataacc
     3421 taacagacta atcgaccgag gcattcatac tcacccgaac cagcctaaac gagataaccc
     3481 ccacagttgg ccgttaggtt agagtcttgt tcggttcttg ggttcttcag cttttgtttt
     3541 tccgctgccc gtcattgtct tctctttctt tgtgctccaa acctttcgcc agggcctgga
     3601 cggatgccgt attcaaacag gttcatgtat ggaaatgact ttggatacgg ccttggatac
     3661 gaccttggat acgacgtcgg ctgcgattac gtgcagaggg agagctacat gttccatgtg
     3721 cgatgccgtg tgagaggtct gatcgcgtgc attcgcggcc gcatttaaat ccccacttca
     3781 gaagttccta tactttctag agaataggaa cttcactata gagtcgaata agggcgacac
     3841 cccataatta gcccgggcga aaggcccagt ctttcgactg agcctttcgt tttatttgat
     3901 gcctggcagt tccctactct cgcatgggga gtccccacac taccatcggc gctacggcgt
     3961 ttcacttctg agttcggcat ggggtcaggt gggaccaccg cgctactgcc gccaggcaaa
     4021 caaggggtgt tatgagccat attcaggtat aaatgggctc gcgataatgt tcagaattgg
     4081 ttaattggtt gtaacactga cccctatttg tttatttttc taaatacatt caaatatgta
     4141 tccgctcatg agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagaatat
     4201 gagtattcaa catttccgtg tcgcccttat tccctttttt gcggcatttt gccttcctgt
     4261 ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg
     4321 agtgggttac atcgaactgg atctcaacag cggtaagatc cttgagagtt ttcgccccga
     4381 agaacgtttt ccaatgatga gcacttttaa agttctgcta tgtggcgcgg tattatcccg
     4441 tattgacgcc gggcaagagc aactcggtcg ccgcatacac tattctcaga atgacttggt
     4501 tgagtactca ccagtcacag aaaagcatct tacggatggc atgacagtaa gagaattatg
     4561 cagtgctgcc ataaccatga gtgataacac tgcggccaac ttacttctga caacgatcgg
     4621 aggaccgaag gagctaaccg cttttttgca caacatgggg gatcatgtaa ctcgccttga
     4681 tcgttgggaa ccggagctga atgaagccat accaaacgac gagcgtgaca ccacgatgcc
     4741 tgtagcgatg gcaacaacgt tgcgcaaact attaactggc gaactactta ctctagcttc
     4801 ccggcaacaa ttaatagact ggatggaggc ggataaagtt gcaggaccac ttctgcgctc
     4861 ggcccttccg gctggctggt ttattgctga taaatccgga gccggtgagc gtggttctcg
     4921 cggtatcatc gcagcgctgg ggccagatgg taagccctcc cgtatcgtag ttatctacac
     4981 gacggggagt caggcaacta tggatgaacg aaatagacag atcgctgaga taggtgcctc
     5041 actgattaag cattggtaag cagagcatta cgctgacttg acgggacggc gcaagctcat
     5101 gaccaaaatc ccttaacgtg agttacgcgc gcgtcgttcc actgagcgtc agaccccgta
     5161 gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa
     5221 acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt
     5281 tttccgaagg taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag
     5341 ccgtagttag cccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta
     5401 atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca
     5461 agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag
     5521 cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa
     5581 agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga
     5641 acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc
     5701 gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc
     5761 ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt
     5821 gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccttt
     5881 gagtgagctg ataccgctcg ccgcagccga acgaccgagc gcagcgagtc agtgagcgag
     5941 gaagcggaag gcgagagtag ggaactgcca ggcatcaaac taagcagaag gcccctgacg
     6001 gatggccttt ttgcgtttct acaaactctt tctgtgttgt aaaacgacgg ccagtcttaa
     6061 gctcgggccc cctgggcggt tctgataacg agtaatcgtt aatccgcaaa taacgtaaaa
     6121 acccgcttcg gcgggttttt ttatgggggg agtttaggga aagagcattt gtcagaatat
     6181 ttaagggcgc ctgtcacttt gcttgatata tgagaattat ttaaccttat aaatgagaaa
     6241 aaagcaacgc actttaaata agatacgttg ctttttcgat tgatgaacac ctataattaa
     6301 actattcatc tattatttat gattttttgt atatacaata tttctagttt gttaaagaga
     6361 attaagaaaa taaatctcga aaataataaa gggaaaatca gtttttgata tcaaaattat
     6421 acatgtcaac gataatacaa aatataatac aaactataag atgttatcag tatttattat
     6481 gcatttagaa taaattttgt gtcgccctta ttcgactcac tatagaagtt cctattctct
     6541 agaaagtata ggaacttcac ttcattttat ttaaatttgc ggccgc
//
