LOCUS       sequence-13758-e        6776 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6776)
  AUTHORS   Garcia-Pedrajas MD, Nadal M, Kapa LB, Perlin MH, Andrews DL, Gold SE
  TITLE     DelsGate, a robust and rapid gene deletion construction method.
  JOURNAL   Fungal Genet Biol. 2008 Apr . 45(4):379-88.
  PUBMED    18248826
REFERENCE   2  (bases 1 to 6776)
  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: "Fungal 
            Genet Biol. 2008 Apr . 45(4):379-88."
COMMENT     Sequence Label: pDONRCBX
FEATURES             Location/Qualifiers
     source          1..6776
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(73..100)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(192..278)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     protein_bind    332..563
                     /gene="mutant version of attP"
                     /label=attP1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             complement(959..1264)
                     /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="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     complement(1061..1080)
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             complement(1606..2265)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     2180..2199
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(2266..2368)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     protein_bind    complement(2513..2744)
                     /gene="mutant version of attP"
                     /label=attP2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(2857..2876)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             2868..3677
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(2935..2954)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      3820..4408
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4309..4328
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgctagcc aggaagagtt
       61 tgtagaaacg caaaaaggcc atccgtcagg atggccttct gcttagtttg atgcctggca
      121 gtttatggcg ggcgtcctgc ccgccaccct ccgggccgtt gcttcacaac gttcaaatcc
      181 gctcccggcg gatttgtcct actcaggaga gcgttcaccg acaaacaaca gataaaacga
      241 aaggcccagt cttccgactg agcctttcgt tttatttgat gcctggcagt tccctactct
      301 cgcgttaacg ctagcatgga tctcgggccc caaataatga ttttattttg actgatagtg
      361 acctgttcgt tgcaacaaat tgatgagcaa tgctttttta taatgccaac tttgtacaaa
      421 aaagctgaac gagaaacgta aaatgatata aatatcaata tattaaatta gattttgcat
      481 aaaaaacaga ctacataata ctgtaaaaca caacatatcc agtcactatg aatcaactac
      541 ttagatggta ttagtgacct gtagtcgacc gacagccttc caaatgttct tcgggtgatg
      601 ctgccaactt agtcgaccga cagccttcca aatgttcttc tcaaacggaa tcgtcgtatc
      661 cagcctactc gctattgtcc tcaatgccgt attaaatcat aaaaagaaat aagaaaaaga
      721 ggtgcgagcc tcttttttgt gtgacaaaat aaaaacatct acctattcat atacgctagt
      781 gtcatagtcc tgaaaatcat ctgcatcaag aacaatttca caactcttat acttttctct
      841 tacaagtcgt tcggcttcat ctggattttc agcctctata cttactaaac gtgataaagt
      901 ttctgtaatt tctactgtat cgacctgcag actggctgtg tataagggag cctgacattt
      961 atattcccca gaacatcagg ttaatggcgt ttttgatgtc attttcgcgg tggctgagat
     1021 cagccacttc ttccccgata acggagaccg gcacactggc catatcggtg gtcatcatgc
     1081 gccagctttc atccccgata tgcaccaccg ggtaaagttc acgggagact ttatctgaca
     1141 gcagacgtgc actggccagg gggatcacca tccgtcgccc gggcgtgtca ataatatcac
     1201 tctgtacatc cacaaacaga cgataacggc tctctctttt ataggtgtaa accttaaact
     1261 gcatttcacc agtccctgtt ctcgtcagca aaagagccgt tcatttcaat aaaccgggcg
     1321 acctcagcca tcccttcctg attttccgct ttccagcgtt cggcacgcag acgacgggct
     1381 tcattctgca tggttgtgct taccagaccg gagatattga catcatatat gccttgagca
     1441 actgatagct gtcgctgtca actgtcactg taatacgctg cttcatagca cacctctttt
     1501 tgacatactt cgggtataca tatcagtata tattcttata ccgcaaaaat cagcgcgcaa
     1561 atacgcatac tgttatctgg cttttagtaa gccggatcca cgcgattacg ccccgccctg
     1621 ccactcatcg cagtactgtt gtaattcatt aagcattctg ccgacatgga agccatcaca
     1681 gacggcatga tgaacctgaa tcgccagcgg catcagcacc ttgtcgcctt gcgtataata
     1741 tttgcccatg gtgaaaacgg gggcgaagaa gttgtccata ttggccacgt ttaaatcaaa
     1801 actggtgaaa ctcacccagg gattggctga gacgaaaaac atattctcaa taaacccttt
     1861 agggaaatag gccaggtttt caccgtaaca cgccacatct tgcgaatata tgtgtagaaa
     1921 ctgccggaaa tcgtcgtggt attcactcca gagcgatgaa aacgtttcag tttgctcatg
     1981 gaaaacggtg taacaagggt gaacactatc ccatatcacc agctcaccgt ctttcattgc
     2041 catacggaat tccggatgag cattcatcag gcgggcaaga atgtgaataa aggccggata
     2101 aaacttgtgc ttatttttct ttacggtctt taaaaaggcc gtaatatcca gctgaacggt
     2161 ctggttatag gtacattgag caactgactg aaatgcctca aaatgttctt tacgatgcca
     2221 ttgggatata tcaacggtgg tatatccagt gatttttttc tccattttag cttccttagc
     2281 tcctgaaaat ctcgataact caaaaaatac gcccggtagt gatcttattt cattatggtg
     2341 aaagttggaa cctcttacgt gccgatcaac gtctcatttt cgccaaaagt tggcccaggg
     2401 cttcccggta tcaacaggga caccaggatt tatttattct gcgaagtgat cttccgtcac
     2461 aggtatttat tcggcgcaaa gtgcgtcggg tgatgctgcc aacttagtcg actacaggtc
     2521 actaatacca tctaagtagt tgattcatag tgactggata tgttgtgttt tacagtatta
     2581 tgtagtctgt tttttatgca aaatctaatt taatatattg atatttatat cattttacgt
     2641 ttctcgttca gctttcttgt acaaagttgg cattataaga aagcattgct tatcaatttg
     2701 ttgcaacgaa caggtcacta tcagtcaaaa taaaatcatt atttgccatc cagctgcagc
     2761 tctggcccgt gtctcaaaat ctctgatgtt acattgcaca agataaaaat atatcatcat
     2821 gaacaataaa actgtctgct tacataaaca gtaatacaag gggtgttatg agccatattc
     2881 aacgggaaac gtcgaggccg cgattaaatt ccaacatgga tgctgattta tatgggtata
     2941 aatgggctcg cgataatgtc gggcaatcag gtgcgacaat ctatcgcttg tatgggaagc
     3001 ccgatgcgcc agagttgttt ctgaaacatg gcaaaggtag cgttgccaat gatgttacag
     3061 atgagatggt cagactaaac tggctgacgg aatttatgcc tcttccgacc atcaagcatt
     3121 ttatccgtac tcctgatgat gcatggttac tcaccactgc gatccccgga aaaacagcat
     3181 tccaggtatt agaagaatat cctgattcag gtgaaaatat tgttgatgcg ctggcagtgt
     3241 tcctgcgccg gttgcattcg attcctgttt gtaattgtcc ttttaacagc gatcgcgtat
     3301 ttcgtctcgc tcaggcgcaa tcacgaatga ataacggttt ggttgatgcg agtgattttg
     3361 atgacgagcg taatggctgg cctgttgaac aagtctggaa agaaatgcat aaacttttgc
     3421 cattctcacc ggattcagtc gtcactcatg gtgatttctc acttgataac cttatttttg
     3481 acgaggggaa attaataggt tgtattgatg ttggacgagt cggaatcgca gaccgatacc
     3541 aggatcttgc catcctatgg aactgcctcg gtgagttttc tccttcatta cagaaacggc
     3601 tttttcaaaa atatggtatt gataatcctg atatgaataa attgcagttt catttgatgc
     3661 tcgatgagtt tttctaatca gaattggtta attggttgta acactggcag agcattacgc
     3721 tgacttgacg ggacggcgca agctcatgac caaaatccct taacgtgagt tttcgttcca
     3781 ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg
     3841 cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga
     3901 tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa
     3961 tactgtcctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc
     4021 tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg
     4081 tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac
     4141 ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct
     4201 acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc
     4261 ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg
     4321 gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg
     4381 ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct
     4441 ggccttttgc tggccttttg ctcactccct gctttgtatg aagaacgccg ttcgctgtcg
     4501 tcaggttgtg gtcgacgcct cttgctgtct ctgccttggt ctgtgcggtg tcgatgtgag
     4561 ctggacccca actgtggact gtccccttca gcttctctat gcctgcttga cttgtgacca
     4621 tgcctgttct cacgtgcaac tctatcgcgg tcttgcctgt gtgaatgtga ctttctctct
     4681 cgacgctcat agctatgacg gtccctgtct cgtctgctgc tatgtgacgg acttgattcg
     4741 cgaagcggcg atgaggacct cctgtctttt cggcaagact cttcgccctt cgagctggtt
     4801 gcctgcatcc tcgactcgga catgatgatg ttgatatcga gcacgttgat gggatcgatc
     4861 ctagaatatc aatcgagcca ccgtcaattt cgagcgctaa gtggagttgt ccgagtcccg
     4921 aatcacaaga attaggctcg tgctctgtga gatctctcgc gaaacccaga tgaaggaaaa
     4981 aaatccggaa gatcgcggaa gaaggtgggg ttcgcatggt ctaacattgt cgcattcctc
     5041 acagtttcgg ctggaaacgg cagggacaat cacgagaaat gcgcgacgag tattcagttg
     5101 tccaaaatta tgtcagcttg aaagttggaa acggcggaag aaaattcgtc aggagggcgt
     5161 tgcggctagg gtgaagaacg gagacaattt cggctttgaa attggctcga ctctgattgg
     5221 atcatgacgc actgctggtc cacaaccgtg tcgaaggtgc cgtctttact acaggtccgc
     5281 tggaagcaaa atggaaaaag ccgctggagc ccgactacag agccgccgtg ttttggtaat
     5341 cagtcggcaa ataggtcagc acagcgcagc gtgacaggtt cttgcaattt acagcacagc
     5401 tcgtccgtct acgactttgc acaccacaaa gtgtgcgggg agcaaaggag ccgatcttgg
     5461 tcgcgcgcaa agccaaggag tcttgaacct gagagtgtgc gtgtcttgtg acgcttgccc
     5521 ttctgtactt tgctgtgaca ctaccaccac atctgtcttg gctttttgtt catacatcca
     5581 caccgaccat gtcgctattc aacgtcagca acggtcttcg taccgctctc cgaccttctg
     5641 ttgccagctc ttcgcgcgtt gctgcctttt ccacaaccgc cgctgcccgt ctcgccacac
     5701 ccacctctga caacgttggc agttcgggca agcctcagca cttgaagcag ttcaagatct
     5761 accgatggaa ccctgacaag ccctcggaga agcctcgtct gcagtcgtac acactggacc
     5821 tcaaccagac cggtccaatg gttctcgacg cgctcatcaa aatcaagaac gaaattgacc
     5881 ctacgctcac cttccgtcgc tcgtgccgtg agggtatctg cggttcgtgc gctatgaata
     5941 ttgacggtgt caacaccctc gcctgcctct gccggatcga caagcagaat gacaccaaga
     6001 tctacccctt gccgcacatg tacattgtca aggacctcgt gccagacttg acccagttct
     6061 acaagcagta ccgatccatc gagcctttcc tcaagtccaa caacacccct tctgagggtg
     6121 aacatcttca gtcgcccgag gagcgtcgtc gactcgacgg tctgtacgag tgcattctgt
     6181 gcgcgtgctg ctccacatcc tgcccctctt actggtggaa tcaggacgag taccttggcc
     6241 ccgccgtgct catgcaggcg taccgatgga tggccgactc gcgtgacgac tttggtgagg
     6301 agcgaagaca gaagctcgag aacacctttt cgctctaccg atgccttacc atcatgaact
     6361 gctccaggac ctgccccaag aacctcaacc ctggtaaggc aattgcacag atcaagaagg
     6421 acatggccgt cggcgcaccc aaggcttccg agcgccctat catggcttcg tcgtaatctt
     6481 gatatatcat atcgttcttt cctcagcact tcttttgtca atttcaaaag tatctaattg
     6541 cattcaactc cgcttgtggt ttgttgttca gtgagagtgg aaacgctacg ggcaagatga
     6601 gggcagtgtt ctggcgacgg aaaagtgtgc aagtgtctgg cctgcgtcct cgctggttcc
     6661 agcagccgat gcaggacgtg tacctagcga tttcttcgac agcctattgt ggcagccgcg
     6721 attcgccaca atcgtacgaa agcgagacga gttgagcgaa gatccccggg cgagtt
//
