LOCUS       sequence-386100-        3786 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  bleomycin (BLE) resistance genes codon-optimized for R. toruloides 
            and expressed under pXYL and tGPD.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3786)
  AUTHORS   Bonturi N, Pinheiro MJ, de Oliveira PM, Rusadze E, Eichinger T, 
            Liudziute G, De Biaggi JS, Brauer A, Remm M, Miranda EA, 
            Ledesma-Amaro R, Lahtvee PJ
  TITLE     Development of a dedicated Golden Gate Assembly Platform (RtGGA) for
            Rhodotorula toruloides.
  JOURNAL   Metab Eng Commun. 2022 May 23;15:e00200. doi: 
            10.1016/j.mec.2022.e00200. eCollection 2022 Dec.
  PUBMED    35662893
REFERENCE   2  (bases 1 to 3786)
  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: "Metab Eng 
            Commun."; date: "2022-05-23"; volume: "15:e00200. doi"; pages: " 
            10.1016/j.mec.2022.e00200. eCollection 2022 Dec"
COMMENT     Sequence Label: (B) BLE marker(R) (SBE262)
FEATURES             Location/Qualifiers
     source          1..3786
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(1622..1638)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1622..1638)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1635..1657)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     rep_origin      complement(1810..2398)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1890..1909)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    complement(2515..2539)
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             complement(join(2639..3430,3431..3499))
                     /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     3262..3281
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3500..3604)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3703..3725
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     3717..3734
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     3718..3734
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 tctctgggtc tcgaggttgt ccgtattcta catcgacgaa gcagggcggg gcggacaggc
       61 gtttgtcgtt gcgcgaggag aacaagggcg gtagagttgt tcttccgcca gctttgtcgc
      121 ccacgctcgt ctgcaacgtc cgtcgtcatt ctaccctcca ctttgacggc cttctcaagc
      181 gcgaggggca cgctccgacg ccggggtgga gcagggaggg gtcgctggag ctcgctcgaa
      241 atggtggacc gtgcgctgcc cgcccaggag agacggaaga gacaggaagc gccggtagaa
      301 acccgggcgg aggaggaaga ggaggttgca gaggaaaccc ggtctgtccc gctcgctggc
      361 tgaaaaagtg cactggggct gcttgatcga tgaggaggag acgcaggctg gacaggggtc
      421 cgacgaagga agcgagccga cgccatgcgg gccgacagac gcttgtccac ttcgctggcg
      481 tctctctctc gctccttctc cctcccagac tcagccaaac tcagtctgtg agtagctttc
      541 gacgcatgcg agcgacagca ctcacccagg ctcgctcact gacccgcttc actcctgccc
      601 gtcctccccc cgcttcgcac cgctgcacgt catctcctcg cgtccccggt tcttcctctc
      661 gcccgtgcag gtgcgtggca tcgtatacac tgttttgcta atttctcgag gacttcagct
      721 aacccgcgta cacttacccg cacgcagaat acacgccatg tcgccacatg gccaagctta
      781 cctcggccgt cccagttctt accgcccgcg acgtcgctgg tgccgtcgaa ttttggaccg
      841 accgactcct tttctcgcga gacttcgttg aggacgactt cgcaggagtc gttcgggacg
      901 acgtcactct cttcatctct gccgttcaag accaggtcgt cccagacaac acccttgcct
      961 gggtttgggt ccgcggtctt gacgaactct acgccgagtg gtcggaagtc gtctcgacca
     1021 actttcgcga cgcatctggc cctgccatga ccgagatcgg cgaacagcct tggggccgcg
     1081 aattcgccct tcgcgaccct gccggaaact gcgtccactt tgttgccgag gaacaagact
     1141 aacgagctcc ccggatccat caccaagggc gtcaacttta tccagcagcc caagctctga
     1201 ctcgctgacc cggctacggg taccgcaaag ttgtagttag cttggcttcc tgtctctctc
     1261 ttcgtcgcgc tcgagtggtt ttcggtgcaa aaggacgtcc gaatcagttc gtagtatgct
     1321 cgtcgcactc gagaggatgc tggcttgcgc cgatgagcaa cgggaacgaa ggcggaggca
     1381 gacttaggct agcgcggacg aggggaagcg agaggcagac gtctatctct cgactccgtc
     1441 atctctcccg catcgtcaac gcacaggctg tgaactgaat tttgtgcatt cactcaaact
     1501 gcgcagtgga agaagcgaag aggctaggcg agaggggcgg gagtctaagt gtggagaccc
     1561 agagaaggct aggtggaggc tcagtgatga taagtctgcg atggtggatg catgtgtcat
     1621 ggtcatagct gtttcctgtg tgaaattgtt atccgctcag agggcacaat cctattccgc
     1681 gctatccgac aatctccaag acattaggtg gagttcagtt cggcgtatgg catatgtcgc
     1741 tggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt
     1801 gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag
     1861 tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc
     1921 cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc
     1981 ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt
     2041 cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt
     2101 atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc
     2161 agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa
     2221 gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa
     2281 gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg
     2341 tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga
     2401 agatcctttg atcttttcta cggggtctga cgctctattc aacaaagccg ccgtcccgtc
     2461 aagtcagcgt aaatgggtag ggggcttcaa atcgtcctcg tgataccaat tcggagcctg
     2521 cttttttgta caaacttgtt gataatggca attcaaggat cttcacctag atccttttaa
     2581 attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt
     2641 accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag
     2701 ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca
     2761 gtgctgcaat gataccgcga gagccacgct caccggctcc agatttatca gcaataaacc
     2821 agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt
     2881 ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg
     2941 ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca
     3001 gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg
     3061 ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca
     3121 tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg
     3181 tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct
     3241 cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca
     3301 tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca
     3361 gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg
     3421 tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac
     3481 ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt
     3541 attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc
     3601 cgcgcacatt tccccgaaaa gtgccagata cctgaaacaa aacccatcgt acggccaagg
     3661 aagtctccaa taactgtgat ccaccacaag cgccagggtt ttcccagtca cgacgttgta
     3721 aaacgacggc cagtcatgca taatccgcac gcatctggaa taaggaagtg ccattccgcc
     3781 tgacct
//
