LOCUS       sequence-420445-        7457 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Gateway cloning compatible vector for transient expression in 
            protoplasts. Arabidopsis UBQ10 promoter for N-terminal mStayGold 
            fusion protein..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7457)
  AUTHORS   Allen JR, Flanagan M, Pathak S, Emenecker SK, Emenecker RJ, Strader 
            LC
  TITLE     An improved toolkit of gateway- and gibson assembly-compatible 
            vectors for protoplast transfection and agrobacterium-mediated plant
            transformation.
  JOURNAL   BMC Res Notes. 2025 Apr 8;18(1):149. doi: 
            10.1186/s13104-025-07231-1.
  PUBMED    40200324
REFERENCE   2  (bases 1 to 7457)
  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: "BMC Res 
            Notes."; date: "2025-04-8"; volume: "18(1):149. doi"; pages: " 
            10.1186/s13104-025-07231-1"
COMMENT     Sequence Label: pLCS110
FEATURES             Location/Qualifiers
     source          1..7457
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(30..52)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     152..171
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     365..387
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     379..396
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     380..396
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             1990..2640
                     /codon_start=1
                     /product="bright and highly photostable monomeric green 
                     fluorescent protein (Ivorra-Molla et al., 2023)"
                     /label=mStayGold (E138D)
                     /note="contains the monomerizing E138D mutation"
                     /translation="MASTPFKFQLKGTINGKSFTVEGEGEGNSHEGSHKGKYVCTSGKL
                     PMSWAALGTSFGYGMKYYTKYPSGLKNWFHEVMPEGFTYDRHIQYKGDGSIHAKHQHFM
                     KNGTYHNIVEFTGQDFKENSPVLTGDMNVSLPNDVQHIPRDDGVECPVTLLYPLLSDKS
                     KCVEAHQNTICKPLHNQPAPDVPYHWIRKQYTQSKDDTEERDHICQSETLEAHL"
     protein_bind    2710..2834
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        join(2859..2864,2865..2882,2883..2889)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             2943..3602
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(3009..3028)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             3944..4249
                     /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     4128..4147
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    complement(4290..4414)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     terminator      4473..5180
                     /label=OCS terminator
                     /note="octopine synthase terminator"
     primer_bind     complement(5237..5253)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5237..5253)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5250..5272)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5261..5277
                     /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)."
     promoter        complement(join(5285..5291,5292..5309,5310..5315))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5330..5351
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5468..5485)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5639..6227)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5719..5738)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6398..7189,7190..7258))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     7021..7040
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7259..7363)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7431..7449
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 ctcgcgcgtt tcggtgatga cggtgaaaac ctctgacaca tgcagctccc ggagacggtc
       61 acagcttgtc tgtaagcgga tgccgggagc agacaagccc gtcagggcgc gtcagcgggt
      121 gttggcgggt gtcggggctg gcttaactat gcggcatcag agcagattgt actgagagtg
      181 caccatatgc ggtgtgaaat accgcacaga tgcgtaagga gaaaataccg catcaggcgc
      241 cattcgccat tcaggctgcg caactgttgg gaagggcgat cggtgcgggc ctcttcgcta
      301 ttacgccagc tggcgaaagg gggatgtgct gcaaggcgat taagttgggt aacgccaggg
      361 ttttcccagt cacgacgttg taaaacgacg gccagtgcca aactttcgcg aagcacccac
      421 cctaagccac attttaatcc ttctgttggc catgccccat caaagttgca cttaacccaa
      481 gattgtggtg gagcttccca tgtttctcgt ctgtcccgac ggtgttgtgg ttggtgcttt
      541 ccttacattc tgagcctctt tccttctaat ccactcatct gcatcttctt gtgtccttac
      601 taatacctca ttggttccaa attccctccc tttaagcacc agctcgtttc tgttcttcca
      661 cagcctccca agtatccaag ggactaaagc ctccacattc ttcagatcag gatattcttg
      721 tttaagatgt tgaactctat ggaggtttgt atgaactgat gatctaggac cggataagtt
      781 cccttcttca tagcgaactt attcaaagaa tgttttgtgt atcattcttg ttacattgtt
      841 attaatgaaa aaatattatt ggtcattgga ctgaacacga gtgttaaata tggaccaggc
      901 cccaaataag atccattgat atatgaatta aataacaaga ataaatcgag tcaccaaacc
      961 acttgccttt tttaacgaga cttgttcacc aacttgatac aaaagtcatt atcctatgca
     1021 aatcaataat catacaaaaa tatccaataa cactaaaaaa ttaaaagaaa tggataattt
     1081 cacaatatgt tatacgataa agaagttact tttccaagaa attcactgat tttataagcc
     1141 cacttgcatt agataaatgg caaaaaaaaa caaaaaggaa aagaaataaa gcacgaagaa
     1201 ttctagaaaa tacgaaatac gcttcaatgc agtgggaccc acggttcaat tattgccaat
     1261 tttcagctcc accgtatatt taaaaaataa aacgataatg ctaaaaaaat ataaatcgta
     1321 acgatcgtta aatctcaacg gctggatctt atgacgaccg ttagaaattg tggttgtcga
     1381 cgagtcagta ataaacggcg tcaaagtggt tgcagccggc acacacgagt cgtgtttatc
     1441 aactcaaagc acaaatactt ttcctcaacc taaaaataag gcaattagcc aaaaacaact
     1501 ttgcgtgtaa acaacgctca atacacgtgt cattttatta ttagctattg cttcaccgcc
     1561 ttagctttct cgtgacctag tcgtcctcgt cttttcttct tcttcttcta taaaacaata
     1621 cccaaagagc tcttcttctt cacaattcag atttcaattt ctcaaaatct taaaaacttt
     1681 ctctcaattc tctctaccgt gatcaaggta aatttctgtg ttccttattc tctcaaaatc
     1741 ttcgattttg ttttcgttcg atcccaattt cgtatatgtt ctttggttta gattctgtta
     1801 atcttagatc gaagacgatt ttctgggttt gatcgttaga tatcatctta attctcgatt
     1861 agggtttcat agatatcatc cgatttgttc aaataatttg agttttgtcg aataattact
     1921 cttcgatttg tgatttctat ctagatctgg tgttagtttc tagtttgtgc gatcgaattt
     1981 gtcctcgaga tggcttctac tccttttaaa tttcagttga agggtactat taatggtaag
     2041 agcttcactg ttgagggaga gggtgaagga aactctcatg agggttcaca taaaggaaag
     2101 tatgtgtgta catctggaaa gttgcctatg tcatgggctg ccctcggtac ttctttcgga
     2161 tatggaatga aatattatac aaaatatcct tctggtctta agaattggtt ccatgaagtg
     2221 atgcctgagg gattcactta cgataggcat attcagtaca agggagatgg ttccattcat
     2281 gctaagcatc agcatttcat gaaaaacgga acttaccata atatcgtgga gttcacaggc
     2341 caagatttta aggaaaactc tcctgttttg actggtgata tgaatgtttc tctcccaaac
     2401 gatgttcaac atattccaag agatgatgga gttgaatgcc ctgttaccct cttgtatcct
     2461 cttctttcag ataaatctaa gtgtgttgag gctcatcaaa acactatctg taagcctttg
     2521 cataatcagc cagctccaga tgtgccatat cattggatca gaaagcaata cacacaatct
     2581 aaggatgata ctgaagaacg tgatcacatc tgtcagtctg aaacacttga agctcatttg
     2641 tccggactca gatcacgagc tcaagcttca aattctgcag tcgacggtac cgcgggcccg
     2701 ggatcatcaa caagtttgta caaaaaagct gaacgagaaa cgtaaaatga tataaatatc
     2761 aatatattaa attagatttt gcataaaaaa cagactacat aatactgtaa aacacaacat
     2821 atccagtcac tatggcggcc gcattaggca ccccaggctt tacactttat gcttccggct
     2881 cgtataatgt gtggattttg agttaggatc cgtcgagatt ttcaggagct aaggaagcta
     2941 aaatggagaa aaaaatcact ggatatacca ccgttgatat atcccaatgg catcgtaaag
     3001 aacattttga ggcatttcag tcagttgctc aatgtaccta taaccagacc gttcagctgg
     3061 atattacggc ctttttaaag accgtaaaga aaaataagca caagttttat ccggccttta
     3121 ttcacattct tgcccgcctg atgaatgctc atccggagtt ccgtatggca atgaaagacg
     3181 gtgagctggt gatatgggat agtgttcacc cttgttacac cgttttccat gagcaaactg
     3241 aaacgttttc atcgctctgg agtgaatacc acgacgattt ccggcagttt ctacacatat
     3301 attcgcaaga tgtggcgtgt tacggtgaaa acctggccta tttccctaaa gggtttattg
     3361 agaatatgtt tttcgtctca gccaatccct gggtgagttt caccagtttt gatttaaacg
     3421 tggccaatat ggacaacttc ttcgcccccg ttttcaccat gggcaaatat tatacgcaag
     3481 gcgacaaggt gctgatgccg ctggcgattc aggttcatca tgccgtttgt gatggcttcc
     3541 atgtcggcag aatgcttaat gaattacaac agtactgcga tgagtggcag ggcggggcgt
     3601 aaagatctgg atccggctta ctaaaagcca gataacagta tgcgtatttg cgcgctgatt
     3661 tttgcggtat aagaatatat actgatatgt atacccgaag tatgtcaaaa agaggtatgc
     3721 tatgaagcag cgtattacag tgacagttga cagcgacagc tatcagttgc tcaaggcata
     3781 tatgatgtca atatctccgg tctggtaagc acaaccatgc agaatgaagc ccgtcgtctg
     3841 cgtgccgaac gctggaaagc ggaaaatcag gaagggatgg ctgaggtcgc ccggtttatt
     3901 gaaatgaacg gctcttttgc tgacgagaac aggggctggt gaaatgcagt ttaaggttta
     3961 cacctataaa agagagagcc gttatcgtct gtttgtggat gtacagagtg atattattga
     4021 cacgcccggg cgacggatgg tgatccccct ggccagtgca cgtctgctgt cagataaagt
     4081 ctcccgtgaa ctttacccgg tggtgcatat cggggatgaa agctggcgca tgatgaccac
     4141 cgatatggcc agtgtgccgg tctccgttat cggggaagaa gtggctgatc tcagccaccg
     4201 cgaaaatgac atcaaaaacg ccattaacct gatgttctgg ggaatataaa tgtcaggctc
     4261 ccttatacac agccagtctg caggtcgacc atagtgactg gatatgttgt gttttacagt
     4321 attatgtagt ctgtttttta tgcaaaatct aatttaatat attgatattt atatcatttt
     4381 acgtttctcg ttcagctttc ttgtacaaag tggttgatga tcctaggtga gtctagagag
     4441 ttaattaaga cccgggacta gtccctagag tcctgcttta atgagatatg cgagacgcct
     4501 atgatcgcat gatatttgct ttcaattctg ttgtgcacgt tgtaaaaaac ctgagcatgt
     4561 gtagctcaga tccttaccgc cggtttcggt tcattctaat gaatatatca cccgttacta
     4621 tcgtattttt atgaataata ttctccgttc aatttactga ttgtacccta ctacttatat
     4681 gtacaatatt aaaatgaaaa caatatattg tgctgaatag gtttatagcg acatctatga
     4741 tagagcgcca caataacaaa caattgcgtt ttattattac aaatccaatt ttaaaaaaag
     4801 cggcagaacc ggtcaaacct aaaagactga ttacataaat cttattcaaa tttcaaaagt
     4861 gccccagggg ctagtatcta cgacacaccg agcggcgaac taataacgct cactgaaggg
     4921 aactccggtt ccccgccggc gcgcatgggt gagattcctt gaagttgagt attggccgtc
     4981 cgctctaccg aaagttacgg gcaccattca acccggtcca gcacggcggc cgggtaaccg
     5041 acttgctgcc ccgagaatta tgcagcattt ttttggtgta tgtgggcccc aaatgaagtg
     5101 caggtcaaac cttgacagtg acgacaaatc gttgggcggg tccagggcga attttgcgac
     5161 aacatgtcga ggctcagcag gacctgcagg tcgactctag aggatccccg ggtaccgagc
     5221 tcgaattcgt aatcatgtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc
     5281 cacacaacat acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct
     5341 aactcacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc
     5401 agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt
     5461 ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag
     5521 ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca
     5581 tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt
     5641 tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc
     5701 gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct
     5761 ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg
     5821 tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca
     5881 agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact
     5941 atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta
     6001 acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta
     6061 actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct
     6121 tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt
     6181 tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga
     6241 tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca
     6301 tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat
     6361 caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg
     6421 cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt
     6481 agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag
     6541 acccacgctc accggctcca gatttatcag caataaacca gccagccgga agggccgagc
     6601 gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag
     6661 ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca
     6721 tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa
     6781 ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga
     6841 tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata
     6901 attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca
     6961 agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg
     7021 ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg
     7081 ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg
     7141 cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag
     7201 gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac
     7261 tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca
     7321 tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag
     7381 tgccacctga cgtctaagaa accattatta tcatgacatt aacctataaa aataggcgta
     7441 tcacgaggcc ctttcgt
//
