LOCUS       sequence-435426-        7135 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  The base plasmid of TUNEYALI for TF57.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7135)
  TITLE     Regulating transcription factors in Yarrowia lipolytica
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7135)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pCfb13247
FEATURES             Location/Qualifiers
     source          1..7135
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_RNA        380..454
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     protein_bind    complement(881..914)
                     /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(1121..1140)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     primer_bind     complement(1146..1165)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        1207..1298
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1299..1367,1368..2159)
                     /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(1517..1536)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2281..2869
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2770..2789
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3023..3040
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     3138..3155
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     3139..3155
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    3625..3658
                     /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)"
     CDS             4800..5828
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     CDS             4800..5828
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     primer_bind     complement(5845..5863)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
ORIGIN
        1 tttatgttaa ctactgtgag gcattaaata gagcttgata tataaaatgt tacatttcac
       61 agtctgaact tttgcagatt acctaatttg gtaagatatt aattatgaac tgaaagttga
      121 tggcatccct aaatttgatg caacggaatg cgtgcgatag tgaatcattg ctaacagatc
      181 aagtaatcgt atcaatcgtt ccacgtctta tccagactcg atttccgccc taacctaagg
      241 aaactgctga gtaatttcga ggttggttcc taacaaaagc caagaaacgc gccgatggtt
      301 tagtggtaaa atccatcgtt gccatcgatg ggcccccggt tcgattccgg gtcggcgcaa
      361 gtttttgaaa ccctgcttgg ttttagagct agaaatagca agttaaaata aggctagtcc
      421 gttatcaact tgaaaaagtg gcaccgagtc ggtggtgctt tttttgtgtg ggtacggtga
      481 aagaatcacg tgaatccttg aacacgtgtc attccaatgg tactgctctc cagtttccag
      541 ctccaatatt gacaccaact cgtcgactat taaagttgtg tggagagagg ttgcatgagt
      601 catatccaat ggccgaggaa gatgggattc tggggtacat gactgaagag ctcttcaatg
      661 tcggtcaaac gtgaatcttc agatagtccc ggggtccaga caaaggacaa gcggcttgca
      721 tgtgtcgaat gtcggcagca aaaggtcaaa tgcgacggcc aggagggctc cgactgctct
      781 cgatgcaagc gaaagggcat tccttgcgtg gtgcaaaaga tcgcgtgcat tcgcggccgc
      841 aatcgcgtgc attcgcggcc gcatttaaat ccccacttca gaagttccta tactttctag
      901 agaataggaa cttcactata gagtcgaata agggcgacac cccataatta gcccgggcga
      961 aaggcccagt ctttcgactg agcctttcgt tttatttgat gcctggcagt tccctactct
     1021 cgcatgggga gtccccacac taccatcggc gctacggcgt ttcacttctg agttcggcat
     1081 ggggtcaggt gggaccaccg cgctactgcc gccaggcaaa caaggggtgt tatgagccat
     1141 attcaggtat aaatgggctc gcgataatgt tcagaattgg ttaattggtt gtaacactga
     1201 cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac
     1261 cctgataaat gcttcaataa tattgaaaaa ggaagaatat gagtattcaa catttccgtg
     1321 tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc
     1381 tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg
     1441 atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga
     1501 gcacttttaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc
     1561 aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag
     1621 aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga
     1681 gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg
     1741 cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga
     1801 atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgtagcgatg gcaacaacgt
     1861 tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact
     1921 ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt
     1981 ttattgctga taaatccgga gccggtgagc gtggttctcg cggtatcatc gcagcgctgg
     2041 ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta
     2101 tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaag
     2161 cagagcatta cgctgacttg acgggacggc gcaagctcat gaccaaaatc ccttaacgtg
     2221 agttacgcgc gcgtcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc
     2281 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
     2341 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
     2401 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag cccaccactt
     2461 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
     2521 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
     2581 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
     2641 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
     2701 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
     2761 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
     2821 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
     2881 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
     2941 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
     3001 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag gcgagagtag
     3061 ggaactgcca ggcatcaaac taagcagaag gcccctgacg gatggccttt ttgcgtttct
     3121 acaaactctt tctgtgttgt aaaacgacgg ccagtcttaa gctcgggccc cctgggcggt
     3181 tctgataacg agtaatcgtt aatccgcaaa taacgtaaaa acccgcttcg gcgggttttt
     3241 ttatgggggg agtttaggga aagagcattt gtcagaatat ttaagggcgc ctgtcacttt
     3301 gcttgatata tgagaattat ttaaccttat aaatgagaaa aaagcaacgc actttaaata
     3361 agatacgttg ctttttcgat tgatgaacac ctataattaa actattcatc tattatttat
     3421 gattttttgt atatacaata tttctagttt gttaaagaga attaagaaaa taaatctcga
     3481 aaataataaa gggaaaatca gtttttgata tcaaaattat acatgtcaac gataatacaa
     3541 aatataatac aaactataag atgttatcag tatttattat gcatttagaa taaattttgt
     3601 gtcgccctta ttcgactcac tatagaagtt cctattctct agaaagtata ggaacttcac
     3661 ttcattttat ttaaatttgc ggccgctatg tctgataaaa ggatgtaaca taggcaagct
     3721 gctcgtgagt gttgagtacg aaccttagat ccaaatcacc cgcacccacg gatatacttg
     3781 cttgaatata cagtagtaag gagtttggcg cccgtttttt cgagccccac acgtttcggt
     3841 gagtatgagc ggcggcagat tcgagcgttt ccggtttccg cggctggacg agagcccatg
     3901 atgggggctc ccaccaccag caatcagggc cctgattaca cacccacctg taatgtcatg
     3961 ctgttcatcg tggttaatgc tgctgtgtgc tgtgtgtgtg tgttgtttgg cgctcattgt
     4021 tgcgttatgc agcgtacacc acaatattgg aagcttatta gcctttctat tttttcgttt
     4081 gcaaggctta acaacattgc tgtggagagg gatggggata tggaggccgc tggagggagt
     4141 cggagaggcg ttttggagcg gcttggcctg gcgcccagct cgcgaaacgc acctaggacc
     4201 ctttggcacg ccgaaatgtg ccacttttca gtctagtaac gccttaccta cgtcattcca
     4261 tgcatgcatg tttgcgcctt ttttcccttg cccttgatcg ccacacagta cagtgcactg
     4321 tacagtggag gttttggggg ggtcttagat gggagctaaa agcggcctag cggtacacta
     4381 gtgggattgt atggagtggc atggagccta ggtggagcct gacaggacgc acgaccggct
     4441 agcccgtgac agacgatggg tggctcctgt tgtccaccgc gtacaaatgt ttgggccaaa
     4501 gtcttgtcag ccttgcttgc gaacctaatt cccaattttg tcacttcgca cccccattga
     4561 tcgagcccta acccctgccc atcaggcaat ccaattaagc tcgcattgtc tgccttgttt
     4621 agtttggctc ctgcccgttt cggcgtccac ttgcacaaac acaaacaagc attatatata
     4681 aggctcgtct ctccctccca accacactca cttttttgcc cgtcttccct tgctaacaca
     4741 aaagtcaaga acacaaacaa ccaccccaac ccccttacac acaagacata tctacagcaa
     4801 tgggtaaaaa gcctgaactc accgcgacgt ctgtcgagaa gtttctgatc gaaaagttcg
     4861 acagcgtctc cgacctgatg cagctctcgg agggcgaaga atctcgtgct ttcagcttcg
     4921 atgtaggagg gcgtggatat gtcctgcggg taaatagctg cgccgatggt ttctacaaag
     4981 atcgttatgt ttatcggcac tttgcatcgg ccgcgctccc gattccggaa gtgcttgaca
     5041 ttggggaatt cagcgagagc ctgacctatt gcatctcccg ccgtgcacag ggtgtcacgt
     5101 tgcaagacct gcctgaaacc gaactgcccg ctgttctgca gccggtcgcg gaggcaatgg
     5161 atgccattgc tgcggccgat cttagccaga cgagcgggtt cggcccattc ggaccgcaag
     5221 gaatcggtca atacactaca tggcgtgatt tcatatgcgc gattgctgat ccccatgtgt
     5281 atcactggca aactgtgatg gacgacaccg tcagtgcgtc cgtcgcgcag gctctcgatg
     5341 agctgatgct ttgggccgag gactgccccg aagtccggca cctcgtgcac gcggatttcg
     5401 gctccaacaa tgtcctgacg gacaatggcc gcataacagc ggtcattgac tggagcgagg
     5461 cgatgttcgg ggattcccaa tacgaggtcg ccaacatctt cttctggagg ccgtggttgg
     5521 cttgtatgga gcagcagacg cgctacttcg agcggaggca tccggagctt gcaggatcgc
     5581 cgcggctccg ggcgtatatg ctccgcattg gtcttgacca actctatcag agcttggttg
     5641 acggcaattt cgatgatgca gcttgggcgc agggtcgatg cgacgcaatc gtccgatccg
     5701 gagccgggac tgtcgggcgt acacaaatcg cccgcagaag cgcggccgtc tggaccgatg
     5761 gctgtgtaga agtactcgcc gatagtggaa accgacgccc cagcactcgt ccgagggcaa
     5821 aggaataatc atgtaattag ttatgtcacg cttacattca cgccctcccc ccacatccgc
     5881 tctaaccgaa aaggaaggag ttagacaacc tgaagtctag gtccctattt atttttttat
     5941 agttatgtta gtattaagaa cgttatttat atttcaaatt tttctttttt ttctgtacag
     6001 acgcgtgtac gcatgtaaca ttatactgaa aaccttgctt gagaaggttt tgggacgctc
     6061 gagcaggctt ggaggcgacg tggcagtggc tcgcgggagg atcggcggga aactccggga
     6121 tatccgtcga gagtttacac gtgaatgggc agcgcaatcc gttgacgacg atacgactgg
     6181 caaagtagcg acgatacctg ccagacaggt gacatgtgca ggccgcacta acaaggaaac
     6241 gggcgctggg gggggcgggc ttctagactt tgcccttgaa caggaatcta gtgggggctt
     6301 gtctttccgc caatggggga gcgcctgttg agcgaccgtg catgctggaa cgccaagtgt
     6361 atgtacagct ggtggtctcg cagcggtatg tgacgggact tacatctctc gttttttcat
     6421 gaccacgttt tcacaggctc ggaggtacgt taaagttttg aaggctgcat ctgaaccgag
     6481 gtatggggga gtttgaacag caacagtgtt ggggctgagg gggccaagat cggggcaagc
     6541 agaggtctta gatcaattgt ggggatccca aagggctcgt tatcaccttt ttccacccaa
     6601 ttcgggtccc aattgatcca ctactggctt gcccaagtta ccccagaaat gccgccccgg
     6661 atttctccaa aaacctaata agcttcatgg aacttggtgg aagtgacttt ctacagagtg
     6721 gagagaaccg tggacacgtg gcaatggcgc tgaccgtgtc cccgagccga atcgacgtga
     6781 ggggagaacg gagtatctgc ggtcatgtga ccttccagag cggcgtcgcc agtgtgcacg
     6841 cggtgacccc cagtttgggt ctctgtcaca cgcatactac ctcggctctc cacatgctga
     6901 actttatctt tcgtggggat cataccgaaa gttgcaacta ccaggtgtat ataaagcctg
     6961 gtagactccc cccactttgg acctcatcca accaagacac acaaaaatgg accgacttaa
     7021 caacctcgcc acccagctcg agcagaaccc cgccaattcg tagataatgg aatacaaatg
     7081 gatatccaga gtatacacat ggatagtata cactgacacg acaattctgt atctc
//
