LOCUS       sequence-386132-        4741 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  integration cassette for ku70 deletion and expression of NAT gene 
            under pXYL for characterization.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4741)
  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 4741)
  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: ku70 insUP+pXYL+NAT+tNOS+ku70 insD (SBE178)
FEATURES             Location/Qualifiers
     source          1..4741
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1284..1856
                     /codon_start=1
                     /gene="Streptomyces noursei nat1"
                     /product="nourseothricin acetyltransferase"
                     /label=NrsR
                     /note="confers resistance to nourseothricin"
                     /translation="MGTTLDDTAYRYRTSVPGDAEAIEALDGSFTTDTVFRATATGDGF
                     TLREVPMDPPLTKVFPDDESDDESDDGEDGDPGSRTFVAYGDDGDLAGFVVVLYSGWNR
                     RLTVEDIEVAPEHRGYGVGRALMGLATEFARERGAGHLWLEVTNVNAPAIHTYRRMGFT
                     LCGLDTALYDGTASDGEQALYMSMPCP"
     primer_bind     complement(2680..2699)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2681..2699)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             complement(2810..3469)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     3384..3403
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(3470..3572)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     rep_origin      3673..4261
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4162..4181
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        4691..4709
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     4691..4708
                     /label=SP6
                     /note="SP6 promoter, forward primer"
ORIGIN
        1 gacttggtac ggagccgcct cctccacctc agcaaccaac ctaccagtcg caccccggat
       61 cgcagtctag tcttccgccg cttcccgctc tcccttcttc ctcgatgccg cctcctccgc
      121 ttcctcggcc caaggcgcct cagagcgccg catcctccgc ctcaacgctt ccgcctccgc
      181 cattctaggc gatgccggga ggaaccgtgc tgaacggaca aggtacggcc ccggcggcag
      241 tccaggcgaa cggggtgcag cagtgaggag cgaacagtat gcatagctgg atcgtcgcat
      301 gcgtcttagt gtcttgcttt cttgttggaa aatcgttgga atcgttggac cttgttgtag
      361 cctctctcac gcaacgactg gcttgtttga cgtctcttca ttgcgtgggg agagtgaagc
      421 gatgtttgca aagtaggaca caacaagcga acggactact gcgaacgtac acaggcgaga
      481 gcgatcgggg gcgaaagtca gtcgtcgtcc tcctcatcgc cgacgatgaa ggttgtccgt
      541 attctacatc gacgaagcag ggcggggcgg acaggcgttt gtcgttgcgc gaggagaaca
      601 agggcggtag agttgttctt ccgccagctt tgtcgcccac gctcgtctgc aacgtccgtc
      661 gtcattctac cctccacttt gacggccttc tcaagcgcga ggggcacgct ccgacgccgg
      721 ggtggagcag ggaggggtcg ctggagctcg ctcgaaatgg tggacagtgc gctgcccgcc
      781 caggagagac ggaagagaca ggaagcgccg gtagaaaccc gggcggagga ggaagaggag
      841 gttgcagagg aaacccggtc tgtcccgctc gctggctgaa aaagtgcact ggggctgctt
      901 gatcgatgag gaggagacgc aggctggaca ggggtccgac gaaggaagcg agccgacgcc
      961 atgcgggccg acagacgctt gtccacttcg ctggcgtctc tctctcgctc cttctccctc
     1021 ccagactcag ccaaactcag tctgtgagta gctttcgacg catgcgagcg acagcactca
     1081 cccaggctcg ctcactgacc cgcttcactc ctgcccgtcc tccccccgct tcgcaccgct
     1141 gcacgtcatc tcctcgcgtc cccggttctt cctctcgccc gtgcaggtgc gtggcatcgt
     1201 atacactgtt ttgctaattt ctcgaggact tcagctaacc cgcgtacact tacccgcacg
     1261 cagaatacac gccatgtcga tcaatgggta ccactcttga cgacacggct taccggtacc
     1321 gcaccagtgt cccgggggac gccgaggcca tcgaggcact ggatgggtcc ttcaccaccg
     1381 acactgtctt ccgcgccacc gccaccgggg acggcttcac cctgcgggag gtgccgatgg
     1441 acccgcccct gaccaaggtg ttccccgacg acgaatcgga cgacgaatcg gacgacgggg
     1501 aggacggcga cccgggctcc cggacgttcg tcgcgtacgg ggacgacggc gacctggcgg
     1561 gcttcgtggt cgtcttgtac tccggctgga accgccggct gaccgtcgag gacatcgagg
     1621 tcgccccgga gcaccggggg tacggggtcg ggcgcgcgtt gatggggctc gcgacggagt
     1681 tcgcccgcga gcggggcgcc gggcacctct ggctggaggt caccaacgtt aacgcaccgg
     1741 ctatccacac gtaccggcgg atggggttca ccctctgcgg cctggacacc gccctgtacg
     1801 acggcaccgc ctcggacggc gagcaggcgc tctacatgag catgccctgc ccctaacgtt
     1861 caaacatttg gcaataaagt ttcttaagat tgaatcctgt tgccggtctt gcgatgatca
     1921 tcatataatt tctgttgaat tacgttaagc atgtaataat taacatgtaa tgcatgacgt
     1981 tatttatgag atgggttttt atgattagag tcccgcaatt atacatttaa tacgcgatag
     2041 aaaacaaaat atagcgcgca aactaggata aattatcgcg cgcggtgtca tctatgttac
     2101 tagatcggga cggatgtggc gaccgggcgc gcgtttcttg ccctcggcgc aagtgggcat
     2161 tcgctcaggc gggcagggtt cttcggcggt agttcacgcg catcagtacg tgccgcgccc
     2221 tttcttcgcc ttacttattg tgagctgacg cactcttgca cagtaacgta caaagcgact
     2281 cggaagacga cgacggcttt gcgcacgtcc acgccaagca cgcctacgag aattttgacg
     2341 aggttgcgcc gcacgagatt gacccgtacg tgccgctgtc tctttcgaat gcatcacaag
     2401 ctcgagctga tgccggccgc aacacccagc ctccgcgatg ttgcggcttc gttcctccac
     2461 gaaatgttcg aggaactctg gcgaattatg aagccgaacg agtatcgcgg cgacaacaaa
     2521 agcgacctcc tcgctcacat ccaccgacac tgcctcccct cgctcctcaa gcacatcccg
     2581 cctcggatcc accacaccgt cgcgccgaaa cggtctaccc attcctgtag tcttcttaat
     2641 taagacgtca gaattctcga ggcggccgca tgtgcgtctc cctatagtga gtcgtattaa
     2701 tttcgcgggc ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct
     2761 catgagtagc accaggcgtt taagggcacc aataactgcc ttaaaaaaat tacgccccgc
     2821 cctgccactc atcgcagtac tgttgtaatt cattaagcat tctgccgaca tggaagccat
     2881 cacaaacggc atgatgaacc tgaatcgcca gcggcatcag caccttgtcg ccttgcgtat
     2941 aatatttgcc catggtgaaa acgggggcga agaagttgtc catattggcc acgtttaaat
     3001 caaaactggt gaaactcacc cagggattgg ctgagacgaa aaacatattc tcaataaacc
     3061 ctttagggaa ataggccagg ttttcaccgt aacacgccac gtcttgcgaa tatatgtgta
     3121 gaaactgccg gaaatcgtcg tggtattcac tccagagcga tgaaaacgtt tcagtttgct
     3181 catggaaaac ggtgtaacaa gggtgaacac tatcccatat caccagctca ccgtctttca
     3241 ttgccatacg gaattccgga tgagcattca tcaggcgggc aagaatgtga ataaaggccg
     3301 gataaaactt gtgcttattt ttctttacgg tctttaaaaa ggccgtaata tccagctgaa
     3361 cggtctggtt ataggtacat tgagcaactg actgaaatgc ctcaaaatgt tctttacgat
     3421 gccattggga tatatcaacg gtggtatatc cagtgatttt tttctccatt ttagcttcct
     3481 tagctcctgg aaatctcgat aactcaaaaa atacgcccgg tagtgatctt atttcattat
     3541 ggtgaaagtt ggaacctctt acgtgccgat caacgtctca ttttcgccaa aagttgtcat
     3601 gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat
     3661 cgaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa
     3721 accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa
     3781 ggtaactggc ttcagcagag cgcagatacc aaatactgtt cttctagtgt agccgtagtt
     3841 aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt
     3901 accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata
     3961 gttgccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt
     4021 ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac
     4081 gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga
     4141 gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg
     4201 ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa
     4261 aaacgccagc aatgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat
     4321 gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcct ttgagtgagc
     4381 tgataccgct cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg aggaagccga
     4441 aaaatcaata atcagacaac aagatgtgcg aactcgatat tttacacgac tctctttacc
     4501 aattctgccc cgaattacac ttaaaacgac tcaacagctt aacgttggct tgccacgcat
     4561 tacttgactg taaaactctc actcttaccg aacttggccg taacctgcca accaaagcga
     4621 gaacaaaaca taacatcaaa cgaatcgacc gattgttagg taatcgtcac ctgcaggaag
     4681 gtttaaacgc atttaggtga cactatagaa gtgtgtatcg ctcgagggat ccgaattcga
     4741 a
//
