LOCUS       sequence-13759-e        5882 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 5882)
  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 5882)
  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: pDONR-A-HYG
FEATURES             Location/Qualifiers
     source          1..5882
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    335..566
                     /gene="mutant version of attP"
                     /label=attP1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             complement(962..1267)
                     /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(1064..1083)
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             complement(1609..2268)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     2183..2202
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(2269..2371)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     protein_bind    complement(2516..2747)
                     /gene="mutant version of attP"
                     /label=attP2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(2860..2879)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             2871..3680
                     /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(2938..2957)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      3823..4411
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4312..4331
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(4546..4573)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(4665..4751)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(4805..5830)
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANILFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
ORIGIN
        1 gaagggcgaa cttaagaagg tatgaccggg tcgtccactt accttgcttg acaaacgcac
       61 caagttatcg tgcaccaagc agcagatgat aataatgtcc tcgttcctgt ctgctaataa
      121 gagtcacact tcgagcgccg ccgctactgc tacaagtggg gctgatctga ccagttgcct
      181 aaatgaacca tcttgtcaaa cgacacaaat tttgtgctca ccgcctggac gactaaacca
      241 aaataggcat tcattgttga cctccactag ctccagccaa gcccaaaaaa tgctccttca
      301 atatcagtta acgctagcat ggatctcggg ccccaaataa tgattttatt ttgactgata
      361 gtgacctgtt cgttgcaaca aattgatgag caatgctttt ttataatgcc aactttgtac
      421 aaaaaagctg aacgagaaac gtaaaatgat ataaatatca atatattaaa ttagattttg
      481 cataaaaaac agactacata atactgtaaa acacaacata tccagtcact atgaatcaac
      541 tacttagatg gtattagtga cctgtagtcg accgacagcc ttccaaatgt tcttcgggtg
      601 atgctgccaa cttagtcgac cgacagcctt ccaaatgttc ttctcaaacg gaatcgtcgt
      661 atccagccta ctcgctattg tcctcaatgc cgtattaaat cataaaaaga aataagaaaa
      721 agaggtgcga gcctcttttt tgtgtgacaa aataaaaaca tctacctatt catatacgct
      781 agtgtcatag tcctgaaaat catctgcatc aagaacaatt tcacaactct tatacttttc
      841 tcttacaagt cgttcggctt catctggatt ttcagcctct atacttacta aacgtgataa
      901 agtttctgta atttctactg tatcgacctg cagactggct gtgtataagg gagcctgaca
      961 tttatattcc ccagaacatc aggttaatgg cgtttttgat gtcattttcg cggtggctga
     1021 gatcagccac ttcttccccg ataacggaga ccggcacact ggccatatcg gtggtcatca
     1081 tgcgccagct ttcatccccg atatgcacca ccgggtaaag ttcacgggag actttatctg
     1141 acagcagacg tgcactggcc agggggatca ccatccgtcg cccgggcgtg tcaataatat
     1201 cactctgtac atccacaaac agacgataac ggctctctct tttataggtg taaaccttaa
     1261 actgcatttc accagtccct gttctcgtca gcaaaagagc cgttcatttc aataaaccgg
     1321 gcgacctcag ccatcccttc ctgattttcc gctttccagc gttcggcacg cagacgacgg
     1381 gcttcattct gcatggttgt gcttaccaga ccggagatat tgacatcata tatgccttga
     1441 gcaactgata gctgtcgctg tcaactgtca ctgtaatacg ctgcttcata gcacacctct
     1501 ttttgacata cttcgggtat acatatcagt atatattctt ataccgcaaa aatcagcgcg
     1561 caaatacgca tactgttatc tggcttttag taagccggat ccacgcgatt acgccccgcc
     1621 ctgccactca tcgcagtact gttgtaattc attaagcatt ctgccgacat ggaagccatc
     1681 acagacggca tgatgaacct gaatcgccag cggcatcagc accttgtcgc cttgcgtata
     1741 atatttgccc atggtgaaaa cgggggcgaa gaagttgtcc atattggcca cgtttaaatc
     1801 aaaactggtg aaactcaccc agggattggc tgagacgaaa aacatattct caataaaccc
     1861 tttagggaaa taggccaggt tttcaccgta acacgccaca tcttgcgaat atatgtgtag
     1921 aaactgccgg aaatcgtcgt ggtattcact ccagagcgat gaaaacgttt cagtttgctc
     1981 atggaaaacg gtgtaacaag ggtgaacact atcccatatc accagctcac cgtctttcat
     2041 tgccatacgg aattccggat gagcattcat caggcgggca agaatgtgaa taaaggccgg
     2101 ataaaacttg tgcttatttt tctttacggt ctttaaaaag gccgtaatat ccagctgaac
     2161 ggtctggtta taggtacatt gagcaactga ctgaaatgcc tcaaaatgtt ctttacgatg
     2221 ccattgggat atatcaacgg tggtatatcc agtgattttt ttctccattt tagcttcctt
     2281 agctcctgaa aatctcgata actcaaaaaa tacgcccggt agtgatctta tttcattatg
     2341 gtgaaagttg gaacctctta cgtgccgatc aacgtctcat tttcgccaaa agttggccca
     2401 gggcttcccg gtatcaacag ggacaccagg atttatttat tctgcgaagt gatcttccgt
     2461 cacaggtatt tattcggcgc aaagtgcgtc gggtgatgct gccaacttag tcgactacag
     2521 gtcactaata ccatctaagt agttgattca tagtgactgg atatgttgtg ttttacagta
     2581 ttatgtagtc tgttttttat gcaaaatcta atttaatata ttgatattta tatcatttta
     2641 cgtttctcgt tcagctttct tgtacaaagt tggcattata agaaagcatt gcttatcaat
     2701 ttgttgcaac gaacaggtca ctatcagtca aaataaaatc attatttgcc atccagctgc
     2761 agctctggcc cgtgtctcaa aatctctgat gttacattgc acaagataaa aatatatcat
     2821 catgaacaat aaaactgtct gcttacataa acagtaatac aaggggtgtt atgagccata
     2881 ttcaacggga aacgtcgagg ccgcgattaa attccaacat ggatgctgat ttatatgggt
     2941 ataaatgggc tcgcgataat gtcgggcaat caggtgcgac aatctatcgc ttgtatggga
     3001 agcccgatgc gccagagttg tttctgaaac atggcaaagg tagcgttgcc aatgatgtta
     3061 cagatgagat ggtcagacta aactggctga cggaatttat gcctcttccg accatcaagc
     3121 attttatccg tactcctgat gatgcatggt tactcaccac tgcgatcccc ggaaaaacag
     3181 cattccaggt attagaagaa tatcctgatt caggtgaaaa tattgttgat gcgctggcag
     3241 tgttcctgcg ccggttgcat tcgattcctg tttgtaattg tccttttaac agcgatcgcg
     3301 tatttcgtct cgctcaggcg caatcacgaa tgaataacgg tttggttgat gcgagtgatt
     3361 ttgatgacga gcgtaatggc tggcctgttg aacaagtctg gaaagaaatg cataaacttt
     3421 tgccattctc accggattca gtcgtcactc atggtgattt ctcacttgat aaccttattt
     3481 ttgacgaggg gaaattaata ggttgtattg atgttggacg agtcggaatc gcagaccgat
     3541 accaggatct tgccatccta tggaactgcc tcggtgagtt ttctccttca ttacagaaac
     3601 ggctttttca aaaatatggt attgataatc ctgatatgaa taaattgcag tttcatttga
     3661 tgctcgatga gtttttctaa tcagaattgg ttaattggtt gtaacactgg cagagcatta
     3721 cgctgacttg acgggacggc gcaagctcat gaccaaaatc ccttaacgtg agttttcgtt
     3781 ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct
     3841 gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc
     3901 ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc
     3961 aaatactgtc cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc
     4021 gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc
     4081 gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg
     4141 aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata
     4201 cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta
     4261 tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc
     4321 ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg
     4381 atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt
     4441 cctggccttt tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt
     4501 ggataaccgt attaccgcta gccaggaaga gtttgtagaa acgcaaaaag gccatccgtc
     4561 aggatggcct tctgcttagt ttgatgcctg gcagtttatg gcgggcgtcc tgcccgccac
     4621 cctccgggcc gttgcttcac aacgttcaaa tccgctcccg gcggatttgt cctactcagg
     4681 agagcgttca ccgacaaaca acagataaaa cgaaaggccc agtcttccga ctgagccttt
     4741 cgttttattt gatgcctggc agttccctac tctcgcgtta actggttccc ggtcggcatc
     4801 tactctattc ctttgccctc ggacgagtgc tggggcgtcg gtttccacta tcggcgagta
     4861 cttctacaca gccatcggtc cagacggccg cgcttctgcg ggcgatttgt gtacgcccga
     4921 cagtcccggc tccggatcgg acgattgcgt cgcatcgacc ctgcgcccaa gctgcatcat
     4981 cgaaattgcc gtcaaccaag ctctgataga gttggtcaag accaatgcgg agcatatacg
     5041 cccggaggcg cggcgatcct gcaagctccg gatgcctccg ctcgaagtag cgcgtctgct
     5101 gctccataca agccaaccac ggcctccaga agaggatgtt ggcgacctcg tattgggaat
     5161 ccccgaacat cgcctcgctc cagtcaatga ccgctgttat gcggccattg tccgtcagga
     5221 cattgttgga gccgaaatcc gcatgcacga ggtgccggac ttcggggcag tcctcggccc
     5281 aaagcatcag ctcatcgaga gcctgcgcga cggacgcact gacggtgtcg tccatcacag
     5341 tttgccagtg atacacatgg ggatcagcaa tcgcgcatat gaaatcacgc catgtagtgt
     5401 attgaccgat tccttgcggt ccgaatgggc cgaacccgct cgtctggcta agatcggccg
     5461 cagcgatcgc atccatggcc tccgcgaccg gctggagaac agcgggcagt tcggtttcag
     5521 gcaggtcttg caacgtgaca ccctgtgcac ggcgggagat gcaataggtc aggctctcgc
     5581 tgaactcccc aatgtcaagc acttccggaa tcgggagcgc ggccgatgca aagtgccgat
     5641 aaacataacg atctttgtag aaaccatcgg cgcagctatt tacccgcagg acatatccac
     5701 gccctcctac atcgaagctg aaagcacgag attcttcgcc ctccgagagc tgcatcaggt
     5761 cggagacgct gtcgaacttt tcgatcagaa acttctcgac agacgtcgcg gtgagttcag
     5821 gctttttcat ttggatgctt gggtagaata ggtaagtcag attgaatctg aaataaaggg
     5881 ag
//
