LOCUS       sequence-361291-        4306 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Transient expression of Citrine in plant cell (Cytosol).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4306)
  AUTHORS   Osaki Y, Kodama Y
  TITLE     Particle bombardment and subcellular protein localization analysis 
            in the aquatic plant Egeria densa.
  JOURNAL   PeerJ. 2017 Sep 7;5:e3779. doi: 10.7717/peerj.3779. eCollection 
            2017.
  PUBMED    28894649
REFERENCE   2  (bases 1 to 4306)
  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: "PeerJ."; 
            date: "2017-09-7"; volume: "5:e3779. doi"; pages: " 
            10.7717/peerj.3779. eCollection 2017"
COMMENT     Sequence Label: pGWT35S-Citrine
FEATURES             Location/Qualifiers
     source          1..4306
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     165..184
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     424..445
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     protein_bind    complement(493..517)
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     misc_feature    561..616
                     /label=TMV Omega
                     /note="translational enhancer from the tobacco mosaic virus
                     5'-leader sequence (Gallie et al., 1988)"
     promoter        complement(636..980)
                     /label=CaMV 35S promoter
                     /note="strong constitutive promoter from cauliflower mosaic
                     virus"
     primer_bind     complement(673..693)
                     /label=35S promoter
                     /note="CaMV 35S promoter, forward primer"
     primer_bind     complement(1056..1075)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1057..1075)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1085..1102)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1085..1101)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1094..1116)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(1310..1329)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1429..1451
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1489..1507)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(1681..1700)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             1692..2507
                     /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="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTSAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(1765..1784)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      2665..3253
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3154..3173
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3407..3424
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    3541..3562
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(3577..3582,3583..3600,3601..3607)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3615..3631
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     3620..3642
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     3639..3655
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3639..3655
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        3673..3691
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     3673..3690
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     terminator      complement(3756..4006)
                     /label=NOS terminator
                     /note="nopaline synthase terminator and poly(A) signal"
     protein_bind    4051..4075
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             complement(join(4080..4306,1..484,485..487,488..490))
                     /codon_start=1
                     /product="enhanced variant of YFP (Heikal et al., 2001)"
                     /label=Citrine
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLMCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(4122..4143)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
ORIGIN
        1 ccttgatgcc gttcttctgc ttgtcggcca tgatatagac gttgtggctg ttgtagttgt
       61 actccagctt gtgccccagg atgttgccgt cctccttgaa gtcgatgccc ttcagctcga
      121 tgcggttcac cagggtgtcg ccctcgaact tcacctcggc gcgggtcttg tagttgccgt
      181 cgtccttgaa gaagatggtg cgctcctgga cgtagccttc gggcatggcg gacttgaaga
      241 agtcgtgctg cttcatgtgg tcggggtagc gggcgaagca catcaggccg tagccgaagg
      301 tggtcacgag ggtgggccag ggcacgggca gcttgccggt ggtgcagatg aacttcaggg
      361 tcagcttgcc gtaggtggca tcgccctcgc cctcgccgga cacgctgaac ttgtggccgt
      421 ttacgtcgcc gtccagctcg accaggatgg gcaccacccc ggtgaacagc tcctcgccct
      481 tgctcaccat ggagcctgct tttttgtaca aacttgtgga gcctcctgag cctccaagct
      541 tggtaccgag ctcgaattct gtaattgtaa atagtaattg taatgttgtt tgttgtttgt
      601 tgttgttggt aattgttgaa ggatctcaag aagcctgtcc tctccaaatg aaatgaactt
      661 ccttatatag aggaagggtc ttgcgaagga tagtgggatt gtgcgtcatc ccttacgtca
      721 gtggagatat cacatcaatc cacttgcttt gaagacgtgg ttggaacgtc ttctttttcc
      781 acgatgttcc tcgtgggtgg gggtccatct ttgggaccac tgtcggtaga ggcatcttga
      841 acgatagcct ttcctttatc gcaatgatgg catttgtagg agccaccttc cttttctact
      901 gtcctttcga taagtgacag atagctgggc aatggaatcc gaggaggttt cccgatatta
      961 ccctttgttg aaaagtctca atagccctcg gtcttctgag cccgggagat ctggcctcga
     1021 gatcccgggt accgagctcg aattctgcgg ccgcgcccta tagtgagtcg tattacacta
     1081 gctcactggc cgtcgtttta caacgtcgtg actgggaaaa ccctggcgtt acccaactta
     1141 atcgccttgc agcacatccc cctttcgcca gctggcgtaa tagcgaagag gcccgcaccg
     1201 atcgcccttc ccaacagttg cgcagcctga atggcgaatg gcgcctgtag taatattttc
     1261 tccttacgca tctgtgcggt atttcacacc gcatatggtg cactctcagt acaatctgct
     1321 ctgatgccgc atagttaagc cagccccgac acccgccaac acccgctgac gcgccctgac
     1381 gggcttgtct gctcccggca tccgcttaca gacaagctgt gaccgtctcc gggagctgca
     1441 tgtgtcagag gttttcaccg tcatcaccga aacgcgcgag acgaaagggc ctcgtgatac
     1501 gcctattttt ataggttaat gtcatgataa taatggtttc ttagacgtca ggtggcactt
     1561 ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt
     1621 atccgctcat gagacaataa ccctgataaa tgcttatgtc tgcttacata aacagtaata
     1681 caaggggtgt tatgagccat attcaacggg aaacgtcttg ctcaaggccg cgattaaatt
     1741 ccaacatgga tgctgattta tatgggtata aatgggctcg cgataatgtc gggcaatcag
     1801 gtgcgacaat ctatcgattg tatgggaagc ccgatgcgcc agagttgttt ctgaaacatg
     1861 gcaaaggtag cgttgccaat gatgttacag atgagatggt cagactaaac tggctgacgg
     1921 aatttatgcc tcttccgacc atcaagcatt ttatccgtac tcctgatgat gcatggttac
     1981 tcacctctgc gatcccaggg aaaacagcat tccaggtatt agaagaatat cctgattcag
     2041 gtgaaaatat tgttgatgcg ctggcagtgt tcctgcgccg gttgcattcg attcctgttt
     2101 gtaattgtcc ttttaacagc gatcgcgtat ttcgtctcgc tcaggcgcaa tcacgaatga
     2161 ataacggttt ggttgatgcg agtgattttg atgacgagcg taatggctgg cctgttgaac
     2221 aagtctggaa agaaatgcat aagctattgc cattctcacc ggattcagtc gtcactcatg
     2281 gtgatttctc acttgataac cttatttttg acgaggggaa attaataggt tgtattgatg
     2341 ttggacgagt cggaatcgca gaccgatacc aggatcttgc catcctatgg aactgcctcg
     2401 gtgagttttc tccttcatta cagaaacggc tttttcaaaa atatggtatt gataatcctg
     2461 atatgaataa attgcagttt catttgatgc tcgatgagtt tttctaatta aaacatatat
     2521 actttagatt gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt
     2581 tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc
     2641 cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt
     2701 gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac
     2761 tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt
     2821 gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct
     2881 gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga
     2941 ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac
     3001 acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg
     3061 agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt
     3121 cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc
     3181 tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg
     3241 gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc
     3301 ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc
     3361 ctttgagtga gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag
     3421 cgaggaagcg gaagagcgcc caatacgcaa accgcctctc cccgcgcgtt ggccgattca
     3481 ttaatgcagc tggcacgaca ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat
     3541 taatgtgagt tagctcactc attaggcacc ccaggcttta cactttatgc ttccggctcg
     3601 tatgttgtgt ggaattgtga gcggataaca atttcacaca ggaaacagct atgaccatga
     3661 ttacgccatg ctatttaggt gacactatag aatacgcggc cgcaagcttg catgcctgca
     3721 ggtcgactct agagcggatc cacaattgcg gccgcatcta gtaacataga tgacaccgcg
     3781 cgcgataatt tatcctagtt tgcgcgctat attttgtttt ctatcgcgta ttaaatgtat
     3841 aattgcggga ctctaatcat aaaaacccat ctcataaata acgtcatgca ttacatgtta
     3901 attattacat gcttaacgta attcaacaga aattatatga taatcatcgc aagaccggca
     3961 acaggattca atcttaagaa actttattgc caaatgtttg aacgattcta gatactagtg
     4021 ctagcccatg gggagcctcc tgagcctcca accactttgt acaagaaagc tgggtctcac
     4081 ttgtacagct cgtccatgcc gagagtgatc ccggcggcgg tcacgaactc cagcaggacc
     4141 atgtgatcgc gcttctcgtt ggggtctttg ctcagggcgg actggtagct caggtagtgg
     4201 ttgtcgggca gcagcacggg gccgtcgccg atgggggtgt tctgctggta gtggtcggcg
     4261 agctgcacgc tgccgtcctc gatgttgtgg cggatcttga agttca
//
