LOCUS       sequence-319282-        3433 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Level 0.5 Golden Gate Vector, loxP-mCHERRY-HDEL-t35s-loxP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3433)
  AUTHORS   Harrington SA, Backhaus AE, Fox S, Rogers C, Borrill P, Uauy C, 
            Richardson A
  TITLE     A heat-shock inducible system for flexible gene expression in 
            cereals.
  JOURNAL   Plant Methods. 2020 Oct 14;16:137. doi: 10.1186/s13007-020-00677-3. 
            eCollection 2020.
  PUBMED    33072173
REFERENCE   2  (bases 1 to 3433)
  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: "Plant 
            Methods."; date: "2020-10-14"; volume: "16:137. doi"; pages: " 
            10.1186/s13007-020-00677-3. eCollection 2020"
COMMENT     Sequence Label: EC71022
FEATURES             Location/Qualifiers
     source          1..3433
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..20
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1..19
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             24..35
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     protein_bind    complement(79..112)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             197..901
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="VSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT
                     QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
                     DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG
                     EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE
                     GRHSTGGMDELYK"
     primer_bind     complement(339..357)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     polyA_signal    957..1131
                     /label=CaMV poly(A) signal
                     /note="cauliflower mosaic virus polyadenylation signal"
     protein_bind    complement(1141..1174)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     complement(1320..1337)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1430..2018)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1510..1529)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(2192..2986)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     promoter        complement(2987..3091)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3392..3414
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     3406..3423
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     3407..3423
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 taatacgact cactataggg cgaattgaag gaaggccgtc aaggccgcat cacttcgtgc
       61 acgtctcata ctgacctaat aacttcgtat agcatacatt atacgaagtt atattaaggg
      121 ttgaatgaaa actaatcttt ttctctttct catcttttca cttctcctat cattatcctc
      181 ggccgaattc ggaggtgtga gcaagggcga ggaggataac atggccatca tcaaggagtt
      241 catgcgcttc aaggtgcaca tggagggctc cgtgaacggc cacgagttcg agatcgaggg
      301 cgagggcgag ggccgcccct acgagggcac ccagaccgcc aagctgaagg tgaccaaggg
      361 tggccccctg cccttcgcct gggacatcct gtcccctcag ttcatgtacg gctccaaggc
      421 ctacgtgaag caccccgccg acatccccga ctacttgaag ctgtccttcc ccgagggctt
      481 caagtgggag cgcgtgatga acttcgagga cggcggcgtg gtgaccgtga cccaggactc
      541 ctccctgcag gacggcgagt tcatctacaa ggtgaagctg cgcggcacca acttcccctc
      601 cgacggcccc gtaatgcaga agaaaaccat gggctgggag gcctcctccg agcggatgta
      661 ccccgaggac ggcgccctga agggcgagat caagcagagg ctgaagctga aggacggcgg
      721 ccactacgac gctgaggtca agaccaccta caaggccaag aagcccgtgc agctgcccgg
      781 cgcctacaac gtcaacatca agttggacat cacctcccac aacgaggact acaccatcgt
      841 ggaacagtac gaacgcgccg agggccgcca ctccaccggc ggcatggacg agctgtacaa
      901 gggtggccat gatgagcttt aagcttctct agctagagtc gatcgacaag ctcgagtttc
      961 tccataataa tgtgtgagta gttcccagat aagggaatta gggttcctat agggtttcgc
     1021 tcatgtgttg agcatataag aaacccttag tatgtatttg tatttgtaaa atacttctat
     1081 caataaaatt tctaattcct aaaaccaaaa tccagtacta aaatccagat cgctgaccta
     1141 ataacttcgt atagcataca ttatacgaag ttatattaag ggttgaatgt gagacgcaca
     1201 gagtgctggg cctcatgggc cttcctttca ctgcccgctt tccagtcggg aaacctgtcg
     1261 tgccagctgc attaacatgg tcatagctgt ttccttgcgt attgggcgct ctccgcttcc
     1321 tcgctcactg actcgctgcg ctcggtcgtt cgggtaaagc ctggggtgcc taatgagcaa
     1381 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc
     1441 tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga
     1501 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc
     1561 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt
     1621 ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct
     1681 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg
     1741 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta
     1801 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct
     1861 acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa
     1921 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt
     1981 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta
     2041 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat
     2101 caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa
     2161 gtatatatga gtaaacttgg tctgacagtt attagaaaaa ttcatccagc agacgataaa
     2221 acgcaatacg ctggctatcc ggtgccgcaa tgccatacag caccagaaaa cgatccgccc
     2281 attcgccgcc cagttcttcc gcaatatcac gggtggccag cgcaatatcc tgataacgat
     2341 ccgccacgcc cagacggccg caatcaataa agccgctaaa acggccattt tccaccataa
     2401 tgttcggcag gcacgcatca ccatgggtca ccaccagatc ttcgccatcc ggcatgctcg
     2461 ctttcagacg cgcaaacagc tctgccggtg ccaggccctg atgttcttca tccagatcat
     2521 cctgatccac caggcccgct tccatacggg tacgcgcacg ttcaatacga tgtttcgcct
     2581 gatgatcaaa cggacaggtc gccgggtcca gggtatgcag acgacgcatg gcatccgcca
     2641 taatgctcac tttttctgcc ggcgccagat ggctagacag cagatcctga cccggcactt
     2701 cgcccagcag cagccaatca cggcccgctt cggtcaccac atccagcacc gccgcacacg
     2761 gaacaccggt ggtggccagc cagctcagac gcgccgcttc atcctgcagc tcgttcagcg
     2821 caccgctcag atcggttttc acaaacagca ccggacgacc ctgcgcgctc agacgaaaca
     2881 ccgccgcatc agagcagcca atggtctgct gcgcccaatc atagccaaac agacgttcca
     2941 cccacgctgc cgggctaccc gcatgcaggc catcctgttc aatcatactc ttcctttttc
     3001 aatattattg aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta
     3061 tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctaaat
     3121 tgtaagcgtt aatattttgt taaaattcgc gttaaatttt tgttaaatca gctcattttt
     3181 taaccaatag gccgaaatcg gcaaaatccc ttataaatca aaagaataga ccgagatagg
     3241 gttgagtggc cgctacaggg cgctcccatt cgccattcag gctgcgcaac tgttgggaag
     3301 ggcgtttcgg tgcgggcctc ttcgctatta cgccagctgg cgaaaggggg atgtgctgca
     3361 aggcgattaa gttgggtaac gccagggttt tcccagtcac gacgttgtaa aacgacggcc
     3421 agtgagcgcg acg
//
