LOCUS       sequence-435956-        7113 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  The base plasmid of TUNEYALI forTF10.
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  JOURNAL   Unpublished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            "Unpublished"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: pCfb13200
FEATURES             Location/Qualifiers
     source          1..7113
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_RNA        308..382
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     protein_bind    complement(787..820)
                     /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(1027..1046)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     primer_bind     complement(1052..1071)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        1113..1204
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1205..1273,1274..2065)
                     /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(1423..1442)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2187..2775
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2676..2695
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2929..2946
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     3044..3061
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     3045..3061
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    3531..3564
                     /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             4706..5734
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     CDS             4706..5734
                     /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(5751..5769)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
ORIGIN
        1 tgcagattac ctaatttggt aagatattaa ttatgaactg aaagttgatg gcatccctaa
       61 atttgatgca acggaatgcg tgcgatagtg aatcattgct aacagatcaa gtaatcgtat
      121 caatcgttcc acgtcttatc cagactcgat ttccgcccta acctaaggaa actgctgagt
      181 aatttcgagg ttggttccta acaaaagcca agaaacgcgc cgatggttta gtggtaaaat
      241 ccatcgttgc catcgatggg cccccggttc gattccgggt cggcgcaaat tttacgtcac
      301 cttcacagtt ttagagctag aaatagcaag ttaaaataag gctagtccgt tatcaacttg
      361 aaaaagtggc accgagtcgg tggtgctttt tttgtgtggg tacggtgata ccttggccac
      421 ggtaatctgt cttgacagcc agcatggcta tgtatcctct cgatcggcac ccacggtagt
      481 cctccagctt gcaaaccacc gttccgatgg cctttttgtc catggacccg tcggcctgct
      541 tctcctcctc gtcttcatca aatgcaagga ctgaagagct cttcaatgtc ggatcgagaa
      601 gagtctggct ctcccgagcc tcggtccaag attgaagaag atctcgagag cgacgaggag
      661 gtgaagacca tcaagagcga agaggtggac aagacacagg acaacgtgga ggacgaggac
      721 gacgaggagg aagaggaaga ggaagagatc gcgtgcattc gcggccgcat ttaaatcccc
      781 acttcagaag ttcctatact ttctagagaa taggaacttc actatagagt cgaataaggg
      841 cgacacccca taattagccc gggcgaaagg cccagtcttt cgactgagcc tttcgtttta
      901 tttgatgcct ggcagttccc tactctcgca tggggagtcc ccacactacc atcggcgcta
      961 cggcgtttca cttctgagtt cggcatgggg tcaggtggga ccaccgcgct actgccgcca
     1021 ggcaaacaag gggtgttatg agccatattc aggtataaat gggctcgcga taatgttcag
     1081 aattggttaa ttggttgtaa cactgacccc tatttgttta tttttctaaa tacattcaaa
     1141 tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa
     1201 gaatatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct
     1261 tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg
     1321 tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg
     1381 ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt
     1441 atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga
     1501 cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga
     1561 attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac
     1621 gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg
     1681 ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac
     1741 gatgcctgta gcgatggcaa caacgttgcg caaactatta actggcgaac tacttactct
     1801 agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct
     1861 gcgctcggcc cttccggctg gctggtttat tgctgataaa tccggagccg gtgagcgtgg
     1921 ttctcgcggt atcatcgcag cgctggggcc agatggtaag ccctcccgta tcgtagttat
     1981 ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg
     2041 tgcctcactg attaagcatt ggtaagcaga gcattacgct gacttgacgg gacggcgcaa
     2101 gctcatgacc aaaatccctt aacgtgagtt acgcgcgcgt cgttccactg agcgtcagac
     2161 cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc
     2221 ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca
     2281 actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta
     2341 gtgtagccgt agttagccca ccacttcaag aactctgtag caccgcctac atacctcgct
     2401 ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg
     2461 gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc
     2521 acacagccca gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta
     2581 tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg
     2641 gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt
     2701 cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg
     2761 cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg
     2821 ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtattacc
     2881 gcctttgagt gagctgatac cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg
     2941 agcgaggaag cggaaggcga gagtagggaa ctgccaggca tcaaactaag cagaaggccc
     3001 ctgacggatg gcctttttgc gtttctacaa actctttctg tgttgtaaaa cgacggccag
     3061 tcttaagctc gggccccctg ggcggttctg ataacgagta atcgttaatc cgcaaataac
     3121 gtaaaaaccc gcttcggcgg gtttttttat ggggggagtt tagggaaaga gcatttgtca
     3181 gaatatttaa gggcgcctgt cactttgctt gatatatgag aattatttaa ccttataaat
     3241 gagaaaaaag caacgcactt taaataagat acgttgcttt ttcgattgat gaacacctat
     3301 aattaaacta ttcatctatt atttatgatt ttttgtatat acaatatttc tagtttgtta
     3361 aagagaatta agaaaataaa tctcgaaaat aataaaggga aaatcagttt ttgatatcaa
     3421 aattatacat gtcaacgata atacaaaata taatacaaac tataagatgt tatcagtatt
     3481 tattatgcat ttagaataaa ttttgtgtcg cccttattcg actcactata gaagttccta
     3541 ttctctagaa agtataggaa cttcacttca ttttatttaa atttgcggcc gctatgtctg
     3601 ataaaaggat gtaacatagg caagctgctc gtgagtgttg agtacgaacc ttagatccaa
     3661 atcacccgca cccacggata tacttgcttg aatatacagt agtaaggagt ttggcgcccg
     3721 ttttttcgag ccccacacgt ttcggtgagt atgagcggcg gcagattcga gcgtttccgg
     3781 tttccgcggc tggacgagag cccatgatgg gggctcccac caccagcaat cagggccctg
     3841 attacacacc cacctgtaat gtcatgctgt tcatcgtggt taatgctgct gtgtgctgtg
     3901 tgtgtgtgtt gtttggcgct cattgttgcg ttatgcagcg tacaccacaa tattggaagc
     3961 ttattagcct ttctattttt tcgtttgcaa ggcttaacaa cattgctgtg gagagggatg
     4021 gggatatgga ggccgctgga gggagtcgga gaggcgtttt ggagcggctt ggcctggcgc
     4081 ccagctcgcg aaacgcacct aggacccttt ggcacgccga aatgtgccac ttttcagtct
     4141 agtaacgcct tacctacgtc attccatgca tgcatgtttg cgcctttttt cccttgccct
     4201 tgatcgccac acagtacagt gcactgtaca gtggaggttt tgggggggtc ttagatggga
     4261 gctaaaagcg gcctagcggt acactagtgg gattgtatgg agtggcatgg agcctaggtg
     4321 gagcctgaca ggacgcacga ccggctagcc cgtgacagac gatgggtggc tcctgttgtc
     4381 caccgcgtac aaatgtttgg gccaaagtct tgtcagcctt gcttgcgaac ctaattccca
     4441 attttgtcac ttcgcacccc cattgatcga gccctaaccc ctgcccatca ggcaatccaa
     4501 ttaagctcgc attgtctgcc ttgtttagtt tggctcctgc ccgtttcggc gtccacttgc
     4561 acaaacacaa acaagcatta tatataaggc tcgtctctcc ctcccaacca cactcacttt
     4621 tttgcccgtc ttcccttgct aacacaaaag tcaagaacac aaacaaccac cccaaccccc
     4681 ttacacacaa gacatatcta cagcaatggg taaaaagcct gaactcaccg cgacgtctgt
     4741 cgagaagttt ctgatcgaaa agttcgacag cgtctccgac ctgatgcagc tctcggaggg
     4801 cgaagaatct cgtgctttca gcttcgatgt aggagggcgt ggatatgtcc tgcgggtaaa
     4861 tagctgcgcc gatggtttct acaaagatcg ttatgtttat cggcactttg catcggccgc
     4921 gctcccgatt ccggaagtgc ttgacattgg ggaattcagc gagagcctga cctattgcat
     4981 ctcccgccgt gcacagggtg tcacgttgca agacctgcct gaaaccgaac tgcccgctgt
     5041 tctgcagccg gtcgcggagg caatggatgc cattgctgcg gccgatctta gccagacgag
     5101 cgggttcggc ccattcggac cgcaaggaat cggtcaatac actacatggc gtgatttcat
     5161 atgcgcgatt gctgatcccc atgtgtatca ctggcaaact gtgatggacg acaccgtcag
     5221 tgcgtccgtc gcgcaggctc tcgatgagct gatgctttgg gccgaggact gccccgaagt
     5281 ccggcacctc gtgcacgcgg atttcggctc caacaatgtc ctgacggaca atggccgcat
     5341 aacagcggtc attgactgga gcgaggcgat gttcggggat tcccaatacg aggtcgccaa
     5401 catcttcttc tggaggccgt ggttggcttg tatggagcag cagacgcgct acttcgagcg
     5461 gaggcatccg gagcttgcag gatcgccgcg gctccgggcg tatatgctcc gcattggtct
     5521 tgaccaactc tatcagagct tggttgacgg caatttcgat gatgcagctt gggcgcaggg
     5581 tcgatgcgac gcaatcgtcc gatccggagc cgggactgtc gggcgtacac aaatcgcccg
     5641 cagaagcgcg gccgtctgga ccgatggctg tgtagaagta ctcgccgata gtggaaaccg
     5701 acgccccagc actcgtccga gggcaaagga ataatcatgt aattagttat gtcacgctta
     5761 cattcacgcc ctccccccac atccgctcta accgaaaagg aaggagttag acaacctgaa
     5821 gtctaggtcc ctatttattt ttttatagtt atgttagtat taagaacgtt atttatattt
     5881 caaatttttc ttttttttct gtacagacgc gtgtacgcat gtaacattat actgaaaacc
     5941 ttgcttgaga aggttttggg acgctcgagc aggcttggag gcgacgtggc agtggctcgc
     6001 gggaggatcg gcgggaaact ccgggatatc cgtcgagagt ttacacgtga atgggcagcg
     6061 caatccgttg acgacgatac gactggcaaa gtagcgacga tacctgccag acaggtgaca
     6121 tgtgcaggcc gcactaacaa ggaaacgggc gctggggggg gcgggcttct agactttgcc
     6181 cttgaacagg aatctagtgg gggcttgtct ttccgccaat gggggagcgc ctgttgagcg
     6241 accgtgcatg ctggaacgcc aagtgtatgt acagctggtg gtctcgcagc ggtatgtgac
     6301 gggacttaca tctctcgttt tttcatgacc acgttttcac aggctcggag gtacgttaaa
     6361 gttttgaagg ctgcatctga accgaggtat gggggagttt gaacagcaac agtgttgggg
     6421 ctgagggggc caagatcggg gcaagcagag gtcttagatc aattgtgggg atcccaaagg
     6481 gctcgttatc acctttttcc acccaattcg ggtcccaatt gatccactac tggcttgccc
     6541 aagttacccc agaaatgccg ccccggattt ctccaaaaac ctaataagct tcatggaact
     6601 tggtggaagt gactttctac agagtggaga gaaccgtgga cacgtggcaa tggcgctgac
     6661 cgtgtccccg agccgaatcg acgtgagggg agaacggagt atctgcggtc atgtgacctt
     6721 ccagagcggc gtcgccagtg tgcacgcggt gacccccagt ttgggtctct gtcacacgca
     6781 tactacctcg gctctccaca tgctgaactt tatctttcgt ggggatcata ccgaaagttg
     6841 caactaccag gtgtatataa agcctggtag actcccccca ctttggacct catccaacca
     6901 agacacacaa aaatggaccg acttaacaac ctcgccaccc agctcgagca gaaccccgcc
     6961 aattcgtaga taatggaata caaatggata tccagagtat acacatggat agtatacact
     7021 gacacgacaa ttctgtatct ctttatgtta actactgtga ggcattaaat agagcttgat
     7081 atataaaatg ttacatttca cagtctgaac ttt
//
