LOCUS       sequence-189234-        6323 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pDONR split marker vector (GenBank accession number: KX904528) for 
            fungal gene deletion. Used with pSM2. .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6323)
  AUTHORS   Rahnama M, Forester N, Ariyawansa KG, Voisey CR, Johnson LJ, Johnson
            RD, Fleetwood DJ
  TITLE     Efficient targeted mutagenesis in Epichloe festucae using a split 
            marker system.
  JOURNAL   J Microbiol Methods. 2016 Dec 22. pii: S0167-7012(16)30368-2. doi: 
            10.1016/j.mimet.2016.12.017.
  PUBMED    28017692
REFERENCE   2  (bases 1 to 6323)
  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: "J Microbiol
            Methods. 2016 Dec 22. pii: S0167-7012(16)30368-2. doi: 
            10.1016/j.mimet.2016.12.017."
COMMENT     Sequence Label: pSM1
FEATURES             Location/Qualifiers
     source          1..6323
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     92..109
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(268..295)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(387..473)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     intron          1133..1248
                     /gene="gpdA"
                     /label=gpdA intron
                     /note="intron from the Aspergillus nidulans 
                     glyceraldehyde-3-phosphate dehydrogenase gene (Punt et al.,
                     1990)"
     primer_bind     2084..2106
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2098..2115
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2099..2115
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    2132..2363
                     /gene="mutant version of attP"
                     /label=attP1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             complement(2759..3064)
                     /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(2861..2880)
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             complement(3384..4067)
                     /codon_start=1
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGRKRR
                     VDPAY"
     primer_bind     3982..4001
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(4068..4170)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     protein_bind    complement(4315..4546)
                     /gene="mutant version of attP"
                     /label=attP2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(4564..4583)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4565..4583)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4588..4604)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4588..4604)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4706..4725)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             4717..5526
                     /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(4784..4803)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      5673..6261
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6162..6181
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga
       61 taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga
      121 gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca
      181 cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaata cgcgtaccgc
      241 tagccaggaa gagtttgtag aaacgcaaaa aggccatccg tcaggatggc cttctgctta
      301 gtttgatgcc tggcagttta tggcgggcgt cctgcccgcc accctccggg ccgttgcttc
      361 acaacgttca aatccgctcc cggcggattt gtcctactca ggagagcgtt caccgacaaa
      421 caacagataa aacgaaaggc ccagtcttcc gactgagcct ttcgttttat ttgatgcctg
      481 gcagttccct actctcgcgt taacgcggcc gccgtcagga cattgttgga gccgaaatcc
      541 gcttgcacga ggtgccggac ttcggggcag tcctcggccc aaagcatcag ctcatcgaga
      601 gcctgcgcga cggacgcact gacggtgtcg tccatcacag tttgccagtg atacacatgg
      661 ggatcagcaa tcgcgcatat gaaatcacgc catgtagtgt attgaccgat tccttgcggt
      721 ccgaatgggc cgaacccgct cgtctggcta agatcggccg cagcgatcgc atccatggcc
      781 tccgcgaccg gctgcagaac agcgggcagt tcggtttcag gcaggtcttg caacgtgaca
      841 ccctgtgcac ggcgggagat gcaataggtc aggctctcgc tgaattcccc aatgtcaagc
      901 acttccggaa tcgggagcgc ggccgatgca aagtgccgat aaacataacg atctttgtag
      961 aaaccatcgg cgcagctatt tacccgcagg acatatccac gccctcctac atcgaagctg
     1021 aaagcacgag attcttcgcc ctccgagagc tgcatcaggt cggagacgct gtcgaacttt
     1081 tcgatcagaa acttctcgac agacgtcgcg gtgagttcag gcatggtgat gtctgctcaa
     1141 gcggggtagc tgttagtcaa gctgcgatga agtgggaaag ctcgaactga aaggttcaaa
     1201 ggaataaggg atgggaagga tggagtatgg atgtagcaaa gtacttactt aggggaaata
     1261 aaggttcttg gatgggaaga tgaatatact gaagatggga aaagaaagag aaaagaaaag
     1321 agcagctggt ggggagagca ggaaaatatg gcaacaaatg ttggactgac gcaacgacct
     1381 tgtcaacccc gccgacacac cgggcggaca gacggggcaa agctgcctac cagggactga
     1441 gggacctcag caggtcgagt gcagagcacc ggatgggtcg actgccagct tgtgttcccg
     1501 gtctgcgccg ctggccagct cctgagcggc ctttccggtt tcatacaccg ggcaaagcag
     1561 gagaggcacg atatttggac gccctacaga tgccggatgg gccaattagg gagcttacgc
     1621 gccgggtact cgctctacct acttcggaga aggtactatc tcgtgaatct tttaccagat
     1681 cggaagcaat tggacttctg tacctaggtt aatggcatgc tatttcgccg acggctatac
     1741 acccctggct tcacattctc cttcgcttac tgccggtgat tcgatgaagc tccatattct
     1801 ccgatgatgc aatagattct tggtcaacga ggggcacacc agcctttcca cttcggggcg
     1861 gaggggcggc cggtcccgga ttaataatca tccactgcac ctcagagccg ccagagctgt
     1921 ctggcgcagt ggcgcttatt actcagccct tctctctgcg tccgtccgtc tctccgcatg
     1981 ccagaaagag tcaccggtca ctgtacagag ctcacgagtt cgtcacattt ttctacaaat
     2041 ggtggaggcg gcggatttta gttaacgcta gcatggatgt tttcccagtc acgacgttgt
     2101 aaaacgacgg ccagtcttaa gctcgggccc caaataatga ttttattttg actgatagtg
     2161 acctgttcgt tgcaacaaat tgatgagcaa tgctttttta taatgccaac tttgtacaaa
     2221 aaagctgaac gagaaacgta aaatgatata aatatcaata tattaaatta gattttgcat
     2281 aaaaaacaga ctacataata ctgtaaaaca caacatatcc agtcactatg aatcaactac
     2341 ttagatggta ttagtgacct gtagtcgacc gacagccttc caaatgttct tcgggtgatg
     2401 ctgccaactt agtcgaccga cagccttcca aatgttcttc tcaaacggaa tcgtcgtatc
     2461 cagcctactc gctattgtcc tcaatgccgt attaaatcat aaaaagaaat aagaaaaaga
     2521 ggtgcgagcc tcttttttgt gtgacaaaat aaaaacatct acctattcat atacgctagt
     2581 gtcatagtcc tgaaaatcat ctgcatcaag aacaatttca caactcttat acttttctct
     2641 tacaagtcgt tcggcttcat ctggattttc agcctctata cttactaaac gtgataaagt
     2701 ttctgtaatt tctactgtat cgacctgcag actggctgtg tataagggag cctgacattt
     2761 atattcccca gaacatcagg ttaatggcgt ttttgatgtc attttcgcgg tggctgagat
     2821 cagccacttc ttccccgata acggagaccg gcacactggc catatcggtg gtcatcatgc
     2881 gccagctttc atccccgata tgcaccaccg ggtaaagttc acgggagact ttatctgaca
     2941 gcagacgtgc actggccagg gggatcacca tccgtcgccc gggcgtgtca ataatatcac
     3001 tctgtacatc cacaaacaga cgataacggc tctctctttt ataggtgtaa accttaaact
     3061 gcatttcacc agcccctgtt ctcgtcagca aaagagccgt tcatttcaat aaaccgggcg
     3121 acctcagcca tcccttcctg attttccgct ttccagcgtt cggcacgcag acgacgggct
     3181 tcattctgca tggttgtgct taccagaccg gagatattga catcatatat gccttgagca
     3241 actgatagct gtcgctgtca actgtcactg taatacgctg cttcatagca tacctctttt
     3301 tgacatactt cgggtataca tatcagtata tattcttata ccgcaaaaat cagcgcgcaa
     3361 atacgcatac tgttatctgg cttttagtaa gccggatcca cgcggcgttt acgccccccc
     3421 tgccactcat cgcagtactg ttgtaattca ttaagcattc tgccgacatg gaagccatca
     3481 caaacggcat gatgaacctg aatcgccagc ggcatcagca ccttgtcgcc ttgcgtataa
     3541 tatttgccca tggtgaaaac gggggcgaag aagttgtcca tattggccac gtttaaatca
     3601 aaactggtga aactcaccca gggattggct gagacgaaaa acatattctc aataaaccct
     3661 ttagggaaat aggccaggtt ttcaccgtaa cacgccacat cttgcgaata tatgtgtaga
     3721 aactgccgga aatcgtcgtg gtattcactc cagagcgatg aaaacgtttc agtttgctca
     3781 tggaaaacgg tgtaacaagg gtgaacacta tcccatatca ccagctcacc gtctttcatt
     3841 gccatacgga attccggatg agcattcatc aggcgggcaa gaatgtgaat aaaggccgga
     3901 taaaacttgt gcttattttt ctttacggtc tttaaaaagg ccgtaatatc cagctgaacg
     3961 gtctggttat aggtacattg agcaactgac tgaaatgcct caaaatgttc tttacgatgc
     4021 cattgggata tatcaacggt ggtatatcca gtgatttttt tctccatttt agcttcctta
     4081 gctcctgaaa atctcgataa ctcaaaaaat acgcccggta gtgatcttat ttcattatgg
     4141 tgaaagttgg aacctcttac gtgccgatca acgtctcatt ttcgccaaaa gttggcccag
     4201 ggcttcccgg tatcaacagg gacaccagga tttatttatt ctgcgaagtg atcttccgtc
     4261 acaggtattt attcggcgca aagtgcgtcg ggtgatgctg ccaacttagt cgactacagg
     4321 tcactaatac catctaagta gttgattcat agtgactgga tatgttgtgt tttacagtat
     4381 tatgtagtct gttttttatg caaaatctaa tttaatatat tgatatttat atcattttac
     4441 gtttctcgtt cagctttctt gtacaaagtt ggcattataa gaaagcattg cttatcaatt
     4501 tgttgcaacg aacaggtcac tatcagtcaa aataaaatca ttatttgcca tccagctgat
     4561 atcccctata gtgagtcgta ttacatggtc atagctgttt cctggcagct ctggcccgtg
     4621 tctcaaaatc tctgatgtta cattgcacaa gataaaaata tatcatcatg aacaataaaa
     4681 ctgtctgctt acataaacag taatacaagg ggtgttatga gccatattca acgggaaacg
     4741 tcgaggccgc gattaaattc caacatggat gctgatttat atgggtataa atgggctcgc
     4801 gataatgtcg ggcaatcagg tgcgacaatc tatcgcttgt atgggaagcc cgatgcgcca
     4861 gagttgtttc tgaaacatgg caaaggtagc gttgccaatg atgttacaga tgagatggtc
     4921 agactaaact ggctgacgga atttatgcct cttccgacca tcaagcattt tatccgtact
     4981 cctgatgatg catggttact caccactgcg atccccggaa aaacagcatt ccaggtatta
     5041 gaagaatatc ctgattcagg tgaaaatatt gttgatgcgc tggcagtgtt cctgcgccgg
     5101 ttgcattcga ttcctgtttg taattgtcct tttaacagcg atcgcgtatt tcgtctcgct
     5161 caggcgcaat cacgaatgaa taacggtttg gttgatgcga gtgattttga tgacgagcgt
     5221 aatggctggc ctgttgaaca agtctggaaa gaaatgcata aacttttgcc attctcaccg
     5281 gattcagtcg tcactcatgg tgatttctca cttgataacc ttatttttga cgaggggaaa
     5341 ttaataggtt gtattgatgt tggacgagtc ggaatcgcag accgatacca ggatcttgcc
     5401 atcctatgga actgcctcgg tgagttttct ccttcattac agaaacggct ttttcaaaaa
     5461 tatggtattg ataatcctga tatgaataaa ttgcagtttc atttgatgct cgatgagttt
     5521 ttctaatcag aattggttaa ttggttgtaa cactggcaga gcattacgct gacttgacgg
     5581 gacggcgcaa gctcatgacc aaaatccctt aacgtgagtt acgcgtcgtt ccactgagcg
     5641 tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc
     5701 tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag
     5761 ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtt
     5821 cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac
     5881 ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc
     5941 gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt
     6001 tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt
     6061 gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc
     6121 ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt
     6181 tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca
     6241 ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt
     6301 tgctggcctt ttgctcacat gtt
//
