LOCUS       sequence-379666-        6985 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Nimble cloning-compatible vector for the analysis of promoter 
            activity in plants.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6985)
  AUTHORS   Yan P, Tuo D, Shen W, Deng H, Zhou P, Gao X
  TITLE     A Nimble Cloning-compatible vector system for high-throughput gene 
            functional analysis in plants.
  JOURNAL   Plant Commun. 2023 Mar 13;4(2):100471. doi: 
            10.1016/j.xplc.2022.100471. Epub 2022 Nov 9.
  PUBMED    36352791
REFERENCE   2  (bases 1 to 6985)
  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 
            Commun."; date: "2023-03-13"; volume: "4(2):100471. doi"; pages: " 
            10.1016/j.xplc.2022.100471. Epub 2022 Nov 9"
COMMENT     Sequence Label: pNC-Green-LUC
FEATURES             Location/Qualifiers
     source          1..6985
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    534..556
                     /label=LB T-DNA repeat
                     /note="left border repeat from nopaline C58 T-DNA 
                     (truncated)"
     promoter        618..963
                     /label=CaMV 35S promoter
                     /note="strong constitutive promoter from cauliflower mosaic
                     virus"
     primer_bind     906..926
                     /label=35S promoter
                     /note="CaMV 35S promoter, forward primer"
     CDS             982..1917
                     /codon_start=1
                     /product="luciferase from the anthozoan coelenterate 
                     Renilla reniformis (sea pansy)"
                     /label=Rluc
                     /translation="MTSKVYDPEQRKRMITGPQWWARCKQMNVLDSFINYYDSEKHAEN
                     AVIFLHGNAASSYLWRHVVPHIEPVARCIIPDLIGMGKSGKSGNGSYRLLDHYKYLTAW
                     FELLNLPKKIIFVGHDWGACLAFHYSYEHQDKIKAIVHAESVVDVIESWDEWPDIEEDI
                     ALIKSEEGEKMVLENNFFVETMLPSKIMRKLEPEEFAAYLEPFKEKGEVRRPTLSWPRE
                     IPLVKGGKPDVVQIVRNYNAYLRASDDLPKMFIESDPGFFSNAIVEGAKKFPNTEFVKV
                     KGLHFSQEDAPDEMGKYIKSFVERVLKNEQ"
     polyA_signal    1972..2148
                     /label=CaMV poly(A) signal
                     /note="cauliflower mosaic virus polyadenylation signal"
     primer_bind     2297..2319
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2311..2328
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2312..2328
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2338..2357
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2338..2356
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        2551..2653
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     CDS             2684..2989
                     /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     2868..2887
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             3077..4729
                     /codon_start=1
                     /gene="luc+"
                     /product="firefly luciferase"
                     /label=luciferase
                     /note="enhanced luc+ version of the luciferase gene"
                     /translation="MEDAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDA
                     HIEVDITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAP
                     ANDIYNERELLNSMGISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQS
                     MYTFVTSHLPPGFNEYDFVPESFDRDKTIALIMNSSGSTGLPKGVALPHRTACVRFSHA
                     RDPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYK
                     IQSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGY
                     GLTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLDTGKTLGVNQRGELCVRGPMIMS
                     GYVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESIL
                     LQHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVV
                     FVDEVPKGLTGKLDARKIREILIKAKKGGKIAV"
     primer_bind     complement(3143..3161)
                     /label=LucNrev
                     /note="Firefly luciferase, reverse primer"
     polyA_signal    4791..4967
                     /label=CaMV poly(A) signal
                     /note="cauliflower mosaic virus polyadenylation signal"
     misc_feature    4977..5001
                     /label=RB T-DNA repeat
                     /note="right border repeat from nopaline C58 T-DNA"
     rep_origin      complement(5092..5680)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5172..5191)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(5851..6666)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(5851..6666)
                     /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
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     6574..6593
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     6658..6677
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      join(6957..6985,1..407)
                     /label=pSa ori
                     /note="origin of replication from bacterial plasmid pSa"
ORIGIN
        1 tttttatccc cggaagcctg tggatagagg gtagttatcc acgtgaaacc gctaatgccc
       61 cgcaaagcct tgattcacgg ggctttccgg cccgctccaa aaactatcca cgtgaaatcg
      121 ctaatcaggg tacgtgaaat cgctaatcgg agtacgtgaa atcgctaata aggtcacgtg
      181 aaatcgctaa tcaaaaaggc acgtgagaac gctaatagcc ctttcagatc aacagcttgc
      241 aaacacccct cgctccggca agtagttaca gcaagtagta tgttcaatta gcttttcaat
      301 tatgaatata tatatcaatt attggtcgcc cttggcttgt ggacaatgcg ctacgcgcac
      361 cggctccgcc cgtggacaac cgcaagcggt tgcccaccgt cgagcgcctt tgcccacaac
      421 ccggcggccg gccgcaacag atcgttttat aaattttttt ttttgaaaaa gaaaaagccc
      481 gaaaggcggc aacctctcgg gcttctggat ttccgatccc cggaattaga tcttggcagg
      541 atatattgtg gtgtaacgtt atcgtacccc tactccaaaa atgtcaaaga tacagtctca
      601 gaagaccaaa gggctattga gacttttcaa caaagggtaa tttcgggaaa cctcctcgga
      661 ttccattgcc cagctatctg tcacttcatc gaaaggacag tagaaaagga aggtggctcc
      721 tacaaatgcc atcattgcga taaaggaaag gctatcattc aagatgcctc tgccgacagt
      781 ggtcccaaag atggaccccc acccacgagg agcatcgtgg aaaaagaaga cgttccaacc
      841 acgtcttcaa agcaagtgga ttgatgtgac atctccactg acgtaaggga tgacgcacaa
      901 tcccactatc cttcgcaaga cccttcctct atataaggaa gttcatttca tttggagagg
      961 acagcccaag ctctagccac catgacttcg aaagtttatg atccagaaca aaggaaacgg
     1021 atgataactg gtccgcagtg gtgggccaga tgtaaacaaa tgaatgttct tgattcattt
     1081 attaattatt atgattcaga aaaacatgca gaaaatgctg ttattttttt acatggtaac
     1141 gcggcctctt cttatttatg gcgacatgtt gtgccacata ttgagccagt agcgcggtgt
     1201 attataccag accttattgg tatgggcaaa tcaggcaaat ctggtaatgg ttcttatagg
     1261 ttacttgatc attacaaata tcttactgca tggtttgaac ttcttaattt accaaagaag
     1321 atcatttttg tcggccatga ttggggtgct tgtttggcat ttcattatag ctatgagcat
     1381 caagataaga tcaaagcaat agttcacgct gaaagtgtag tagatgtgat tgaatcatgg
     1441 gatgaatggc ctgatattga agaagatatt gcgttgatca aatctgaaga aggagaaaaa
     1501 atggttttgg agaataactt cttcgtggaa accatgttgc catcaaaaat catgagaaag
     1561 ttagaaccag aagaatttgc agcatatctt gaaccattca aagagaaagg tgaagttcgt
     1621 cgtccaacat tatcatggcc tcgtgaaatc ccgttagtaa aaggtggtaa acctgacgtt
     1681 gtacaaattg ttaggaatta taatgcttat ctacgtgcaa gtgatgattt accaaaaatg
     1741 tttattgaat cggacccagg attcttttcc aatgctattg ttgaaggtgc caagaagttt
     1801 cctaatactg aatttgtcaa agtaaaaggt cttcattttt cgcaagaaga tgcacctgat
     1861 gaaatgggaa aatatatcaa atcgttcgtt gagcgagttc tcaaaaatga acaataattc
     1921 tagggtacgc tgaaatcacc agtctctctc tacaaatcta tctctctcta ttttctccat
     1981 aaataatgtg tgagtagttt cccgataagg gaaattaggg ttcttatagg gtttcgctca
     2041 tgtgttgagc atataagaaa cccttagtat gtatttgtat ttgtaaaata cttctatcaa
     2101 taaaatttct aattcctaaa accaaaatcc agtactaaaa tccagatcga taacattaac
     2161 gcttacaatt tccattcgcc attcaggctg cgcaactgtt gggaagggcg atcggtgcgg
     2221 gcctcttcgc tattacgcca gctggcgaaa gggggatgtg ctgcaaggcg attaagttgg
     2281 gtaacgccag ggttttccca gtcacgacgt tgtaaaacga cggccagtga gcgcgcgtaa
     2341 tacgactcac tatagggcga attgggtacc agtggtctct gtccagtcct ggcctcgtcg
     2401 gccattctcg actaagttgg cagcatcacc cgacgcactt tgcgccgaat aaatacctgt
     2461 gacggaagat cacttcgcag aataaataaa tcctggtgtc cctgttgata ccgggaagcc
     2521 ctgggccaac ttttggcgaa aatgagacgt tgatcggcac gtaagaggtt ccaactttca
     2581 ccataatgaa ataagatcac taccgggcgt attttttgag ttatcgagat tttcaggagc
     2641 taaggaagct aaacttttgc tgacgagaac agggactggt gaaatgcagt ttaaggttta
     2701 cacctataaa agagagagcc gttatcgtct gtttgtggat gtacagagtg atattattga
     2761 cacgcctggg cgacggatgg tgatccccct ggccagtgca cgtctgctgt cagataaagt
     2821 ctcccgtgaa ctttacccgg tggtgcatat cggggatgaa agctggcgca tgatgaccac
     2881 cgatatggcc agtgtgccgg tatccgttat cggggaagaa gtggctgatc tcagccaccg
     2941 cgaaaatgac atcaaaaacg ccattaacct gatgttctgg ggaatataaa tgtcaggctc
     3001 ccttatacac agggccagtc tggccacttg tggtctgctg agaccgcggc cgccaccgcg
     3061 gtggagatcg aattccatgg aagacgccaa aaacataaag aaaggcccgg cgccattcta
     3121 tccgctggaa gatggaaccg ctggagagca actgcataag gctatgaaga gatacgccct
     3181 ggttcctgga acaattgctt ttacagatgc acatatcgag gtggacatca cttacgctga
     3241 gtacttcgaa atgtccgttc ggttggcaga agctatgaaa cgatatgggc tgaatacaaa
     3301 tcacagaatc gtcgtatgca gtgaaaactc tcttcaattc tttatgccgg tgttgggcgc
     3361 gttatttatc ggagttgcag ttgcgcccgc gaacgacatt tataatgaac gtgaattgct
     3421 caacagtatg ggcatttcgc agcctaccgt ggtgttcgtt tccaaaaagg ggttgcaaaa
     3481 aattttgaac gtgcaaaaaa agctcccaat catccaaaaa attattatca tggattctaa
     3541 aacggattac cagggatttc agtcgatgta cacgttcgtc acatctcatc tacctcccgg
     3601 ttttaatgaa tacgattttg tgccagagtc cttcgatagg gacaagacaa ttgcactgat
     3661 catgaactcc tctggatcta ctggtctgcc taaaggtgtc gctctgcctc atagaactgc
     3721 ctgcgtgaga ttctcgcatg ccagagatcc tatttttggc aatcaaatca ttccggatac
     3781 tgcgatttta agtgttgttc cattccatca cggttttgga atgtttacta cactcggata
     3841 tttgatatgt ggatttcgag tcgtcttaat gtatagattt gaagaagagc tgtttctgag
     3901 gagccttcag gattacaaga ttcaaagtgc gctgctggtg ccaaccctat tctccttctt
     3961 cgccaaaagc actctgattg acaaatacga tttatctaat ttacacgaaa ttgcttctgg
     4021 tggcgctccc ctctctaagg aagtcgggga agcggttgcc aagaggttcc atctgccagg
     4081 tatcaggcaa ggatatgggc tcactgagac tacatcagct attctgatta cacccgaggg
     4141 ggatgataaa ccgggcgcgg tcggtaaagt tgttccattt tttgaagcga aggttgtgga
     4201 tctggatacc gggaaaacgc tgggcgttaa tcaaagaggc gaactgtgtg tgagaggtcc
     4261 tatgattatg tccggttatg taaacaatcc ggaagcgacc aacgccttga ttgacaagga
     4321 tggatggcta cattctggag acatagctta ctgggacgaa gacgaacact tcttcatcgt
     4381 tgaccgcctg aagtctctga ttaagtacaa aggctatcag gtggctcccg ctgaattgga
     4441 atccatcttg ctccaacacc ccaacatctt cgacgcaggt gtcgcaggtc ttcccgacga
     4501 tgacgccggt gaacttcccg ccgccgttgt tgttttggag cacggaaaga cgatgacgga
     4561 aaaagagatc gtggattacg tcgccagtca agtaacaacc gcgaaaaagt tgcgcggagg
     4621 agttgtgttt gtggacgaag taccgaaagg tcttaccgga aaactcgacg caagaaaaat
     4681 cagagagatc ctcataaagg ccaagaaggg cggaaagatc gccgtgtaat tctagagaat
     4741 tcggtacgct gaaatcacca gtctctctct acaaatctat ctctctctat tttctccata
     4801 aataatgtgt gagtagtttc ccgataaggg aaattagggt tcttataggg tttcgctcat
     4861 gtgttgagca tataagaaac ccttagtatg tatttgtatt tgtaaaatac ttctatcaat
     4921 aaaatttcta attcctaaaa ccaaaatcca gtactaaaat ccagatccac tagccttgac
     4981 aggatatatt ggcgggtaaa ctaagtcgct gtatgtgttt gtttgagatc tcatgtgagc
     5041 aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag
     5101 gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc
     5161 gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt
     5221 tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct
     5281 ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg
     5341 ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct
     5401 tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat
     5461 tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg
     5521 ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaag
     5581 aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt
     5641 ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc
     5701 tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt
     5761 atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta
     5821 aagtatatat gtgtaacatt ggtctagtga ttagaaaaac tcatcgagca tcaaatgaaa
     5881 ctgcaattta ttcatatcag gattatcaat accatatttt tgaaaaagcc gtttctgtaa
     5941 tgaaggagaa aactcaccga ggcagttcca taggatggca agatcctggt atcggtctgc
     6001 gattccgact cgtccaacat caatacaacc tattaatttc ccctcgtcaa aaataaggtt
     6061 atcaagtgag aaatcaccat gagtgacgac tgaatccggt gagaatggca aaagtttatg
     6121 catttctttc cagacttgtt caacaggcca gccattacgc tcgtcatcaa aatcactcgc
     6181 atcaaccaaa ccgttattca ttcgtgattg cgcctgagcg agacgaaata cgcgatcgct
     6241 gttaaaagga caattacaaa caggaatcga atgcaaccgg cgcaggaaca ctgccagcgc
     6301 atcaacaata ttttcacctg aatcaggata ttcttctaat acctggaatg ctgttttccc
     6361 tgggatcgca gtggtgagta accatgcatc atcaggagta cggataaaat gcttgatggt
     6421 cggaagaggc ataaattccg tcagccagtt tagtctgacc atctcatctg taacatcatt
     6481 ggcaacgcta cctttgccat gtttcagaaa caactctggc gcatcgggct tcccatacaa
     6541 tcggtagatt gtcgcacctg attgcccgac attatcgcga gcccatttat acccatataa
     6601 atcagcatcc atgttggaat ttaatcgcgg ccttgagcaa gacgtttccc gttgaatatg
     6661 gctcataaca ccccttgtat tactgtttat gtaagcagac agttttattg ttcatgatga
     6721 tatattttta tcttgtgcaa tgtaacatca gagattttga gacacaacgt ggctttgttg
     6781 aataaatcga acttttgctg agttgaagga tcagatcacg catcttcccg acaacgcaga
     6841 ccgttccgtg gcaaagcaaa agttcaaaat caccaactgg tccacctaca acaaagctct
     6901 catcaaccgt ggctccctca ctttctggct ggatgatggg gcgattcagg cgatccccat
     6961 ccaacagccc gccgtcgagc gggct
//
