LOCUS       sequence-229341-        5268 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Part of GoldenGate Yarrowia lipolytica toolkit for assembling an 
            expression plasmid for Y. lipolytic yeast. Destination vector for 
            the insertion of the assembly. It already has some of the parts 
            needed for transformation in Y. lipolytica.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5268)
  AUTHORS   Larroude M, Park YK, Soudier P, Kubiak M, Nicaud JM, Rossignol T
  TITLE     A modular Golden Gate toolkit for Yarrowia lipolytica synthetic 
            biology.
  JOURNAL   Microb Biotechnol. 2019 May 31. doi: 10.1111/1751-7915.13427.
  PUBMED    31148366
REFERENCE   2  (bases 1 to 5268)
  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: "Microb 
            Biotechnol. 2019 May 31. doi: 10.1111/1751-7915.13427."
COMMENT     Sequence Label: GGE114
FEATURES             Location/Qualifiers
     source          1..5268
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    596..617
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(632..637,638..655,656..662)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    670..686
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     RBS             704..715
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             722..1399
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /translation="MASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKM
                     RLKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHS
                     TGA"
     terminator      1444..1515
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      1531..1558
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     terminator      2004..2061
                     /label=his operon terminator
                     /note="This putative transcriptin terminator from the E. 
                     coli his operon has a 2-bp deletion introduced during 
                     synthesis. Its efficiency has not been determined."
     primer_bind     complement(2085..2102)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2256..2844)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2336..2355)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3015..3806,3807..3875))
                     /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     3638..3657
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3876..3980)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4048..4066
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     terminator      complement(4070..4113)
                     /label=bacterial terminator
                     /note="putative bacterial transcription terminator"
     protein_bind    complement(4452..4485)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
ORIGIN
        1 gaaaccgtgc ttaagagcaa gttccttgag ggggagcaca gtgccggcgt aggtgaagtc
       61 gtcaatgatg tcgatatggg tcttgatcat gcacacataa ggtccgacct tatcggcaag
      121 ctcaatgagc tccttggtgg tggtaacatc cagagaagca cacaggttgg ttttcttggc
      181 tgccacgagc ttgagcactc gagcggcaaa ggcggacttg tggacgttag ctcgagcttc
      241 gtaggagggc attttggtgg tgaagaggag actgaaataa atttagtctg cagaactttt
      301 tatcggaacc ttatctgggg cagtgaagta tatgttatgg taatagttac gagttagttg
      361 aacttataga tagactggac tatacggcta tcggtccaaa ttagaaagaa cgtcaatggc
      421 tctctgggcg cgaattcgta taacttcgta tagcaggagt tatccgaagc gaacggtgag
      481 accggatccc tagagcaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat
      541 gcagctggca cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaatg
      601 tgagttagct cactcattag gcaccccagg ctttacactt tatgcttccg gctcgtatgt
      661 tgtgtggaat tgtgagcgga taacaatttc acacatacta gagaaagagg agaaatacta
      721 gatggcttcc tccgaagacg ttatcaaaga gttcatgcgt ttcaaagttc gtatggaagg
      781 ttccgttaac ggtcacgagt tcgaaatcga aggtgaaggt gaaggtcgtc cgtacgaagg
      841 tacccagacc gctaaactga aagttaccaa aggtggtccg ctgccgttcg cttgggacat
      901 cctgtccccg cagttccagt acggttccaa agcttacgtt aaacacccgg ctgacatccc
      961 ggactacctg aaactgtcct tcccggaagg tttcaaatgg gaacgtgtta tgaacttcga
     1021 agacggtggt gttgttaccg ttacccagga ctcctccctg caagacggtg agttcatcta
     1081 caaagttaaa ctgcgtggta ccaacttccc gtccgacggt ccggttatgc agaaaaaaac
     1141 catgggttgg gaagcttcca ccgaacgtat gtacccggaa gacggtgctc tgaaaggtga
     1201 aatcaaaatg cgtctgaaac tgaaagacgg tggtcactac gacgctgaag ttaaaaccac
     1261 ctacatggct aaaaaaccgg ttcagctgcc gggtgcttac aaaaccgaca tcaaactgga
     1321 catcacctcc cacaacgaag actacaccat cgttgaacag tacgaacgtg ctgaaggtcg
     1381 tcactccacc ggtgcttaat aacgctgata gtgctagtgt agatcgctac tagagccagg
     1441 catcaaataa aacgaaaggc tcagtcgaaa gactgggcct ttcgttttat ctgttgtttg
     1501 tcggtgaacg ctctctacta gagtcacact ggctcacctt cgggtgggcc tttctgcgtt
     1561 tatatatgta cagcggtctc tgagtcatgt gtaacactcg ctctggagag ttagtcatcc
     1621 gacagggtaa ctctaatctc ccaacacctt attaactctg cgtaactgta actcttcttg
     1681 ccacgtcgat cttactcaat tttcctgctc atcatctgct ggattgttgt ctatcgtctg
     1741 gctctaatac atttattgtt tattgcccaa acaactttca ttgcacgtaa gtgaattgtt
     1801 ttataacagc gttcgccaat tgctgcgcca tcgtcgtccg gctgtcctac cgttaggggt
     1861 agtgtgtctc acactaccga ggttactaga gttgggaaag cgatactgcc tcggacacac
     1921 cacctgggtc ttacgactgc agagagaatc ggcgttacct ctctcacaaa gcccttcagt
     1981 gcggccgctg cgtcttcctg cagtccggca aaaaagggca aggtgtcacc accctgccct
     2041 ttttctttaa aaccgaaaag attacttcgc gttatgcagg cttcctcgct cactgactcg
     2101 ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg
     2161 ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag
     2221 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc acaggctccg cccccctgac
     2281 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
     2341 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
     2401 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
     2461 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     2521 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
     2581 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
     2641 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca
     2701 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
     2761 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
     2821 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
     2881 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc
     2941 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa
     3001 acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
     3061 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
     3121 ttaccatctg gccccagtgc tgcaatgata ccgcgcgacc cacgctcacc ggctccagat
     3181 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
     3241 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
     3301 aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt
     3361 ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg
     3421 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc
     3481 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc
     3541 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg
     3601 cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga
     3661 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta
     3721 ccgctgttga gatccagttc gatataaccc actcgtgcac ccaactgatc ttcagcatct
     3781 tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag
     3841 ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca atattattga
     3901 agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat
     3961 aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgt ctaagaaacc
     4021 attattatca tgacattaac ctataaaaat aggcgtatca cgaggcagaa tttcagataa
     4081 aaaaaatcct tagctttcgc taaggatgat ttctggaatt cgaagagcct gcggccgctg
     4141 tcgggaaccg cgttcaggtg gaacaggacc acctcccttg cacttcttgg tatatcagta
     4201 taggctgatg tattcatagt ggggtttttc ataataaatt tactaacggc aggcaacatt
     4261 cactcggctt aaacgcaaaa cggaccgtct tgatatcttc tgacgcattg accaccgaga
     4321 aatagtgtta gttaccgggt gagttattgt tcttctacac aggcgacgcc catcgtctag
     4381 agttgatgta ctaactcaga tttcactacc taccctatcc ctggtacgca caaagcactt
     4441 tgctagaagg tataacttcg tatagcatac attatacgaa gttattctga attccgagaa
     4501 acacaacaac atgccccatt ggacagacca tgcggataca caggttgtgc agtaccatac
     4561 atactcgatc agacaggtcg tctgaccatc atacaagctg aacagcgctc catacttgca
     4621 cgctctctat atacacagtt aaattacata tccatagtct aacctctaac agttaatctt
     4681 ctggtaagcc tcccagccag ccttctggta tcgcttggcc tcctcaatag gatctcggtt
     4741 ctggccgtac agacctcggc cgacaattat gatatccgtt ccggtagaca tgacatcctc
     4801 aacagttcgg tactgctgtc cgagagcgtc tcccttgtcg tcaagaccca ccccgggggt
     4861 cagaataagc cagtcctcag agtcgccctt aggtcggttc tgggcaatga agccaaccac
     4921 aaactcgggg tcggatcggg caagctcaat ggtctgcttg gagtactcgc cagtggccag
     4981 agagcccttg caagacagct cggccagcat gagcagacct ctggccagct tctcgttggg
     5041 agaggggact aggaactcct tgtactggga gttctcgtag tcagagacgt cctccttctt
     5101 ctgttcagag acagtttcct cggcaccagc tcgcaggcca gcaatgattc cggttccggg
     5161 tacaccgtgg gcgttggtga tatcggacca ctcggcgatt cggtgacacc ggtactggtg
     5221 cttgacagtg ttgccaatat ctgcgaactt tctgtcctcg aacaggaa
//
