LOCUS       sequence-460318-        4024 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  PrDGA with the overhangs for SapI restriction site can be insert 
            into the base plasmid of TUNEYALI.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4024)
  TITLE     Regulating transcription factors in Yarrowia lipolytica
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 4024)
  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            "Unpublished"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: pCfb12566
FEATURES             Location/Qualifiers
     source          1..4024
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      302..890
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     791..810
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1044..1061
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    1178..1199
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(1214..1219,1220..1237,1238..1244)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1252..1268
                     /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)."
     primer_bind     1257..1279
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     1276..1292
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1276..1292
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        1310..1328
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     1310..1327
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(1983..2002)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1984..2002)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2009..2026)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2009..2025)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2018..2040)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     CDS             2163..2465
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="QFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDKV
                     SRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     2344..2363
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             2814..3608
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             2814..3608
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(2868..2887)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3478..3497
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     regulatory      3808..3817
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(3814..4024,1..164)
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
ORIGIN
        1 tggacgagct gtacgccgag tggtcggagg tcgtgtccac gaacttccgg gacgcctccg
       61 ggccggccat gaccgagatc ggcgagcagc cgtgggggcg ggagttcgcc ctgcgcgacc
      121 cggccggcaa ctgcgtgcac ttcgtggccg aggagcagga ctgacacgtg ctaaaacttc
      181 atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc
      241 cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt
      301 cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac
      361 cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct
      421 tcagcagagc gcagatacca aatactgttc ttctagtgta gccgtagtta ggccaccact
      481 tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg
      541 ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata
      601 aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga
      661 cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag
      721 ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg
      781 agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac
      841 ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca
      901 acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg
      961 cgttatcccc tgattctgtg gataaccgta ttaccgcctt tgagtgagct gataccgctc
     1021 gccgcagccg aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa gagcgcccaa
     1081 tacgcaaacc gcctctcccc gcgcgttggc cgattcatta atgcagctgg cacgacaggt
     1141 ttcccgactg gaaagcgggc agtgagcgca acgcaattaa tgtgagttag ctcactcatt
     1201 aggcacccca ggctttacac tttatgcttc cggctcgtat gttgtgtgga attgtgagcg
     1261 gataacaatt tcacacagga aacagctatg accatgatta cgccaagcta tttaggtgac
     1321 actatagaat actcaagcta tgcatcaagc ttggtaccga gctcggatcc actagtaacg
     1381 gccgccagtg tgcgggaatt cgcccttgct cttcagactt tctcccacca ccaccaacac
     1441 tccctttcca ttcccacacc gttcctctct cgtgcgtcat ccttgcgcaa tcatcttcgt
     1501 ctgcgacata ttgtacgaca tacagtacca cggaacgttt cagaccgtcg acgtgaacac
     1561 atcttaggaa cagcaacctg agctacagaa atctatctat aggcggataa aaaaacgcac
     1621 ccactgctcg tcctccttgc tcctcgaaac cgactcctct acacacgtca aatccgaggt
     1681 tgaaatcttc cccacatttg gcagccaaac cagcacatcc cagcaacctc gcacagcgcc
     1741 gaaatcgacc tgtcgacttg gccacaaaaa aaagcaccgg ctctgcaaca gttctcacga
     1801 ccaattacgt acaagtacga aatcgttcgt ggaccgtgac tgataagctc ccactttttc
     1861 ttctaacaac aggcaacaga caagtcacac aaaacaaaag ctatgagaag agcaagggcg
     1921 aattctgcag atatccatca cactggcggc cgctcgagca tgcatctaga gggcccaatt
     1981 cgccctatag tgagtcgtat tacaattcac tggccgtcgt tttacaacgt cgtgactggg
     2041 aaaaccctgg cgttacccaa cttaatcgcc ttgcagcaca tccccctttc gccagctggc
     2101 gtaatagcga agaggcccgc accgatcgcc cttcccaaca gttgcgcagc ctatacgtac
     2161 ggcagtttaa ggtttacacc tataaaagag agagccgtta tcgtctgttt gtggatgtac
     2221 agagtgatat tattgacacg ccggggcgac ggatggtgat ccccctggcc agtgcacgtc
     2281 tgctgtcaga taaagtctcc cgtgaacttt acccggtggt gcatatcggg gatgaaagct
     2341 ggcgcatgat gaccaccgat atggccagtg tgccggtctc cgttatcggg gaagaagtgg
     2401 ctgatctcag ccaccgcgaa aatgacatca aaaacgccat taacctgatg ttctggggaa
     2461 tataaatgtc aggcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttcacgtaga
     2521 aagccagtcc gcagaaacgg tgctgacccc ggatgaatgt cagctactgg gctatctgga
     2581 caagggaaaa cgcaagcgca aagagaaagc aggtagcttg cagtgggctt acatggcgat
     2641 agctagactg ggcggtttta tggacagcaa gcgaaccgga attgccagct ggggcgccct
     2701 ctggtaaggt tgggaagccc tgcaaagtaa actggatggc tttcttgccg ccaaggatct
     2761 gatggcgcag gggatcaagc tctgatcaag agacaggatg aggatcgttt cgcatgattg
     2821 aacaagatgg attgcacgca ggttctccgg ccgcttgggt ggagaggcta ttcggctatg
     2881 actgggcaca acagacaatc ggctgctctg atgccgccgt gttccggctg tcagcgcagg
     2941 ggcgcccggt tctttttgtc aagaccgacc tgtccggtgc cctgaatgaa ctgcaagacg
     3001 aggcagcgcg gctatcgtgg ctggccacga cgggcgttcc ttgcgcagct gtgctcgacg
     3061 ttgtcactga agcgggaagg gactggctgc tattgggcga agtgccgggg caggatctcc
     3121 tgtcatctca ccttgctcct gccgagaaag tatccatcat ggctgatgca atgcggcggc
     3181 tgcatacgct tgatccggct acctgcccat tcgaccacca agcgaaacat cgcatcgagc
     3241 gagcacgtac tcggatggaa gccggtcttg tcgatcagga tgatctggac gaagagcatc
     3301 aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc gagcatgccc gacggcgagg
     3361 atctcgtcgt gacccatggc gatgcctgct tgccgaatat catggtggaa aatggccgct
     3421 tttctggatt catcgactgt ggccggctgg gtgtggcgga ccgctatcag gacatagcgt
     3481 tggctacccg tgatattgct gaagagcttg gcggcgaatg ggctgaccgc ttcctcgtgc
     3541 tttacggtat cgccgctccc gattcgcagc gcatcgcctt ctatcgcctt cttgacgagt
     3601 tcttctgaat tattaacgct tacaatttcc tgatgcggta ttttctcctt acgcatctgt
     3661 gcggtatttc acaccgcatc aggtggcact tttcggggaa atgtgcgcgg aacccctatt
     3721 tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa
     3781 atgcttcaat aatagcacgt gaggagggcc accatggcca agttgaccag tgccgttccg
     3841 gtgctcaccg cgcgcgacgt cgccggagcg gtcgagttct ggaccgaccg gctcgggttc
     3901 tcccgggact tcgtggagga cgacttcgcc ggtgtggtcc gggacgacgt gaccctgttc
     3961 atcagcgcgg tccaggacca ggtggtgccg gacaacaccc tggcctgggt gtgggtgcgc
     4021 ggcc
//
