LOCUS       sequence-435380-        7113 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  The base plasmid of TUNEYALI for TF29.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7113)
  TITLE     Regulating transcription factors in Yarrowia lipolytica
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7113)
  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            "Unpublished"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: pCfb13219
FEATURES             Location/Qualifiers
     source          1..7113
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_RNA        222..296
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     protein_bind    complement(701..734)
                     /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(941..960)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     primer_bind     complement(966..985)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        1027..1118
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1119..1187,1188..1979)
                     /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(1337..1356)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2101..2689
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2590..2609
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2843..2860
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     2958..2975
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2959..2975
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    3445..3478
                     /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             4620..5648
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     CDS             4620..5648
                     /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(5665..5683)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
ORIGIN
        1 agtgaatcat tgctaacaga tcaagtaatc gtatcaatcg ttccacgtct tatccagact
       61 cgatttccgc cctaacctaa ggaaactgct gagtaatttc gaggttggtt cctaacaaaa
      121 gccaagaaac gcgccgatgg tttagtggta aaatccatcg ttgccatcga tgggcccccg
      181 gttcgattcc gggtcggcgc agggtctgac gatgatgatg cgttttagag ctagaaatag
      241 caagttaaaa taaggctagt ccgttatcaa cttgaaaaag tggcaccgag tcggtggtgc
      301 tttttttgtg tgggtacggt ccaaaagtcg ttggaccacc gcaagactcg gagccgcgga
      361 cacacacaag atgggtcggg tccacggaca cgaacacaca ttgacacgtg ctgaccaaac
      421 cattgattgt gatgactgat gctggtgttg ttgtcgatga atgaaaaacg tgtccgacaa
      481 atgactgaag agctcttcaa tgaagaaggc atacgaactg tctgtgctca ccggcaccca
      541 ggttctgctg ctggtcgtgt ctgagaccgg acttgtgtac accttcacta cccccaagct
      601 gcagcccctt gtcacaaagc ccgagggcaa gaacctgatc caggcctgcc ttaacgcgtc
      661 gatcgcgtgc attcgcggcc gcatttaaat ccccacttca gaagttccta tactttctag
      721 agaataggaa cttcactata gagtcgaata agggcgacac cccataatta gcccgggcga
      781 aaggcccagt ctttcgactg agcctttcgt tttatttgat gcctggcagt tccctactct
      841 cgcatgggga gtccccacac taccatcggc gctacggcgt ttcacttctg agttcggcat
      901 ggggtcaggt gggaccaccg cgctactgcc gccaggcaaa caaggggtgt tatgagccat
      961 attcaggtat aaatgggctc gcgataatgt tcagaattgg ttaattggtt gtaacactga
     1021 cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac
     1081 cctgataaat gcttcaataa tattgaaaaa ggaagaatat gagtattcaa catttccgtg
     1141 tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc
     1201 tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg
     1261 atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga
     1321 gcacttttaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc
     1381 aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag
     1441 aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga
     1501 gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg
     1561 cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga
     1621 atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgtagcgatg gcaacaacgt
     1681 tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact
     1741 ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt
     1801 ttattgctga taaatccgga gccggtgagc gtggttctcg cggtatcatc gcagcgctgg
     1861 ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta
     1921 tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaag
     1981 cagagcatta cgctgacttg acgggacggc gcaagctcat gaccaaaatc ccttaacgtg
     2041 agttacgcgc gcgtcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc
     2101 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
     2161 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
     2221 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag cccaccactt
     2281 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
     2341 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
     2401 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
     2461 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
     2521 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
     2581 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
     2641 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
     2701 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
     2761 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
     2821 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag gcgagagtag
     2881 ggaactgcca ggcatcaaac taagcagaag gcccctgacg gatggccttt ttgcgtttct
     2941 acaaactctt tctgtgttgt aaaacgacgg ccagtcttaa gctcgggccc cctgggcggt
     3001 tctgataacg agtaatcgtt aatccgcaaa taacgtaaaa acccgcttcg gcgggttttt
     3061 ttatgggggg agtttaggga aagagcattt gtcagaatat ttaagggcgc ctgtcacttt
     3121 gcttgatata tgagaattat ttaaccttat aaatgagaaa aaagcaacgc actttaaata
     3181 agatacgttg ctttttcgat tgatgaacac ctataattaa actattcatc tattatttat
     3241 gattttttgt atatacaata tttctagttt gttaaagaga attaagaaaa taaatctcga
     3301 aaataataaa gggaaaatca gtttttgata tcaaaattat acatgtcaac gataatacaa
     3361 aatataatac aaactataag atgttatcag tatttattat gcatttagaa taaattttgt
     3421 gtcgccctta ttcgactcac tatagaagtt cctattctct agaaagtata ggaacttcac
     3481 ttcattttat ttaaatttgc ggccgctatg tctgataaaa ggatgtaaca taggcaagct
     3541 gctcgtgagt gttgagtacg aaccttagat ccaaatcacc cgcacccacg gatatacttg
     3601 cttgaatata cagtagtaag gagtttggcg cccgtttttt cgagccccac acgtttcggt
     3661 gagtatgagc ggcggcagat tcgagcgttt ccggtttccg cggctggacg agagcccatg
     3721 atgggggctc ccaccaccag caatcagggc cctgattaca cacccacctg taatgtcatg
     3781 ctgttcatcg tggttaatgc tgctgtgtgc tgtgtgtgtg tgttgtttgg cgctcattgt
     3841 tgcgttatgc agcgtacacc acaatattgg aagcttatta gcctttctat tttttcgttt
     3901 gcaaggctta acaacattgc tgtggagagg gatggggata tggaggccgc tggagggagt
     3961 cggagaggcg ttttggagcg gcttggcctg gcgcccagct cgcgaaacgc acctaggacc
     4021 ctttggcacg ccgaaatgtg ccacttttca gtctagtaac gccttaccta cgtcattcca
     4081 tgcatgcatg tttgcgcctt ttttcccttg cccttgatcg ccacacagta cagtgcactg
     4141 tacagtggag gttttggggg ggtcttagat gggagctaaa agcggcctag cggtacacta
     4201 gtgggattgt atggagtggc atggagccta ggtggagcct gacaggacgc acgaccggct
     4261 agcccgtgac agacgatggg tggctcctgt tgtccaccgc gtacaaatgt ttgggccaaa
     4321 gtcttgtcag ccttgcttgc gaacctaatt cccaattttg tcacttcgca cccccattga
     4381 tcgagcccta acccctgccc atcaggcaat ccaattaagc tcgcattgtc tgccttgttt
     4441 agtttggctc ctgcccgttt cggcgtccac ttgcacaaac acaaacaagc attatatata
     4501 aggctcgtct ctccctccca accacactca cttttttgcc cgtcttccct tgctaacaca
     4561 aaagtcaaga acacaaacaa ccaccccaac ccccttacac acaagacata tctacagcaa
     4621 tgggtaaaaa gcctgaactc accgcgacgt ctgtcgagaa gtttctgatc gaaaagttcg
     4681 acagcgtctc cgacctgatg cagctctcgg agggcgaaga atctcgtgct ttcagcttcg
     4741 atgtaggagg gcgtggatat gtcctgcggg taaatagctg cgccgatggt ttctacaaag
     4801 atcgttatgt ttatcggcac tttgcatcgg ccgcgctccc gattccggaa gtgcttgaca
     4861 ttggggaatt cagcgagagc ctgacctatt gcatctcccg ccgtgcacag ggtgtcacgt
     4921 tgcaagacct gcctgaaacc gaactgcccg ctgttctgca gccggtcgcg gaggcaatgg
     4981 atgccattgc tgcggccgat cttagccaga cgagcgggtt cggcccattc ggaccgcaag
     5041 gaatcggtca atacactaca tggcgtgatt tcatatgcgc gattgctgat ccccatgtgt
     5101 atcactggca aactgtgatg gacgacaccg tcagtgcgtc cgtcgcgcag gctctcgatg
     5161 agctgatgct ttgggccgag gactgccccg aagtccggca cctcgtgcac gcggatttcg
     5221 gctccaacaa tgtcctgacg gacaatggcc gcataacagc ggtcattgac tggagcgagg
     5281 cgatgttcgg ggattcccaa tacgaggtcg ccaacatctt cttctggagg ccgtggttgg
     5341 cttgtatgga gcagcagacg cgctacttcg agcggaggca tccggagctt gcaggatcgc
     5401 cgcggctccg ggcgtatatg ctccgcattg gtcttgacca actctatcag agcttggttg
     5461 acggcaattt cgatgatgca gcttgggcgc agggtcgatg cgacgcaatc gtccgatccg
     5521 gagccgggac tgtcgggcgt acacaaatcg cccgcagaag cgcggccgtc tggaccgatg
     5581 gctgtgtaga agtactcgcc gatagtggaa accgacgccc cagcactcgt ccgagggcaa
     5641 aggaataatc atgtaattag ttatgtcacg cttacattca cgccctcccc ccacatccgc
     5701 tctaaccgaa aaggaaggag ttagacaacc tgaagtctag gtccctattt atttttttat
     5761 agttatgtta gtattaagaa cgttatttat atttcaaatt tttctttttt ttctgtacag
     5821 acgcgtgtac gcatgtaaca ttatactgaa aaccttgctt gagaaggttt tgggacgctc
     5881 gagcaggctt ggaggcgacg tggcagtggc tcgcgggagg atcggcggga aactccggga
     5941 tatccgtcga gagtttacac gtgaatgggc agcgcaatcc gttgacgacg atacgactgg
     6001 caaagtagcg acgatacctg ccagacaggt gacatgtgca ggccgcacta acaaggaaac
     6061 gggcgctggg gggggcgggc ttctagactt tgcccttgaa caggaatcta gtgggggctt
     6121 gtctttccgc caatggggga gcgcctgttg agcgaccgtg catgctggaa cgccaagtgt
     6181 atgtacagct ggtggtctcg cagcggtatg tgacgggact tacatctctc gttttttcat
     6241 gaccacgttt tcacaggctc ggaggtacgt taaagttttg aaggctgcat ctgaaccgag
     6301 gtatggggga gtttgaacag caacagtgtt ggggctgagg gggccaagat cggggcaagc
     6361 agaggtctta gatcaattgt ggggatccca aagggctcgt tatcaccttt ttccacccaa
     6421 ttcgggtccc aattgatcca ctactggctt gcccaagtta ccccagaaat gccgccccgg
     6481 atttctccaa aaacctaata agcttcatgg aacttggtgg aagtgacttt ctacagagtg
     6541 gagagaaccg tggacacgtg gcaatggcgc tgaccgtgtc cccgagccga atcgacgtga
     6601 ggggagaacg gagtatctgc ggtcatgtga ccttccagag cggcgtcgcc agtgtgcacg
     6661 cggtgacccc cagtttgggt ctctgtcaca cgcatactac ctcggctctc cacatgctga
     6721 actttatctt tcgtggggat cataccgaaa gttgcaacta ccaggtgtat ataaagcctg
     6781 gtagactccc cccactttgg acctcatcca accaagacac acaaaaatgg accgacttaa
     6841 caacctcgcc acccagctcg agcagaaccc cgccaattcg tagataatgg aatacaaatg
     6901 gatatccaga gtatacacat ggatagtata cactgacacg acaattctgt atctctttat
     6961 gttaactact gtgaggcatt aaatagagct tgatatataa aatgttacat ttcacagtct
     7021 gaacttttgc agattaccta atttggtaag atattaatta tgaactgaaa gttgatggca
     7081 tccctaaatt tgatgcaacg gaatgcgtgc gat
//
