LOCUS pGreen_dualluc_ORF_sensor 7072 bp DNA circular SYN 26-AUG-2024
DEFINITION Dual-luciferase reporter for microRNA targeting in plants.
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 7072)
AUTHORS Liu Q, Wang F, Axtell MJ
TITLE Analysis of complementarity requirements for plant MicroRNA
targeting using a Nicotiana benthamiana quantitative transient
assay.
JOURNAL Plant Cell. 2014 Feb;26(2):741-53. doi: 10.1105/tpc.113.120972. Epub
2014 Feb 7.
PUBMED 24510721
REFERENCE 2 (bases 1 to 7072)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Aug 26, 2024 from SnapGene Server 7.0.3
https://www.biofeng.com
COMMENT SGRef: number: 1; type: "Journal Article"; doi:
"10.1105/tpc.113.120972"; journalName: "Plant Cell"; date:
"2014-02"; volume: "26"; issue: "2"; pages: "741-53"
FEATURES Location/Qualifiers
source 1..7072
/mol_type="other DNA"
/organism="synthetic DNA construct"
misc_feature 542..564
/label=LB T-DNA repeat
/note="left border repeat from nopaline C58 T-DNA
(truncated)"
terminator complement(587..839)
/label=NOS terminator
/note="nopaline synthase terminator and poly(A) signal"
CDS complement(854..1789)
/codon_start=1
/product="luciferase from the anthozoan coelenterate
Renilla reniformis (sea pansy)"
/label=Rluc
/translation="MTSKVYDPEQRKRMITGPQWWARCKQMNVLDSFINYYDSEKHAEN
AVIFLHGNAASSYLWRHVVPHIEPVARCIIPDLIGMGKSGKSGNGSYRLLDHYKYLTAW
FELLNLPKKIIFVGHDWGACLAFHYSYEHQDKIKAIVHAESVVDVIESWDEWPDIEEDI
ALIKSEEGEKMVLENNFFVETMLPSKIMRKLEPEEFAAYLEPFKEKGEVRRPTLSWPRE
IPLVKGGKPDVVQIVRNYNAYLRASDDLPKMFIESDPGFFSNAIVEGAKKFPNTEFVKV
KGLHFSQEDAPDEMGKYIKSFVERVLKNEQ"
promoter complement(1823..2002)
/label=NOS promoter
/note="nopaline synthase promoter"
primer_bind 2234..2256
/label=M13/pUC Forward
/note="In lacZ gene"
primer_bind 2248..2265
/label=M13 Forward
/note="In lacZ gene. Also called M13-F20 or M13 (-21)
Forward"
primer_bind 2249..2265
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
primer_bind 2275..2294
/label=T7
/note="T7 promoter, forward primer"
promoter 2275..2293
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
promoter 2368..2711
/label=CaMV 35S promoter
/note="strong constitutive promoter from cauliflower mosaic
virus"
primer_bind 2656..2676
/label=35S promoter
/note="CaMV 35S promoter, forward primer"
CDS 2718..4367
/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(2784..2802)
/label=LucNrev
/note="Firefly luciferase, reverse primer"
promoter complement(4671..4689)
/label=T3 promoter
/note="promoter for bacteriophage T3 RNA polymerase"
primer_bind complement(4710..4726)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(4710..4726)
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind complement(4723..4745)
/label=M13/pUC Reverse
/note="In lacZ gene"
protein_bind 4734..4750
/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(4758..4789)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 4804..4825
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
primer_bind complement(4942..4959)
/label=L4440
/note="L4440 vector, forward primer"
misc_feature 5064..5088
/label=RB T-DNA repeat
/note="right border repeat from nopaline C58 T-DNA"
rep_origin complement(5179..5767)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(5259..5278)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(5938..6753)
/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
KPDAPELFLKHGKGSVANVVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
primer_bind 6661..6680
/label=Kan-R
/note="Kanamycin resistance gene, reverse primer"
primer_bind 6745..6764
/label=pENTR-R
/note="pENTR vectors, reverse primer"
rep_origin join(7044..7072,1..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 cgagcgccag cgcctttgcc
421 cacaacccgg cggccggccg caacagatcg ttttataaat tttttttttt gaaaaagaaa
481 aagcccgaaa ggcggcaacc tctcgggctt ctggatttcc gatccccgga attagagatc
541 ttggcaggat atattgtggt gtaacgtatc agcttgcatg ccggtcgatc tagtaacata
601 gatgacaccg cgcgcgataa tttatcctag tttgcgcgct atattttgtt ttctatcgcg
661 tattaaatgt ataattgcgg gactctaatc ataaaaaccc atctcataaa taacgtcatg
721 cattacatgt taattattac atgcttaacg taattcaaca gaaattatat gataatcatc
781 gcaagaccgg caacaggatt caatcttaag aaactttatt gccaaatgtt tgaacgatct
841 gcttgactct agattattgt tcatttttga gaactcgctc aacgaacgat ttgatatatt
901 ttcccatttc atcaggtgca tcttcttgcg aaaaatgaag accttttact ttgacaaatt
961 cagtattagg aaacttcttg gcaccttcaa caatagcatt ggaaaagaat cctgggtccg
1021 attcaataaa catttttggt aaatcatcac ttgcacgtag ataagcatta taattcctaa
1081 caatttgtac aacgtcaggt ttaccacctt ttactaacgg gatttcacga ggccatgata
1141 atgttggacg acgaacttca cctttctctt tgaatggttc aagatatgct gcaaattctt
1201 ctggttctaa ctttctcatg atttttgatg gcaacatggt ttccacgaag aagttattct
1261 ccaaaaccat tttttctcct tcttcagatt tgatcaacgc aatatcttct tcaatatcag
1321 gccattcatc ccatgattca atcacatcta ctacactttc agcgtgaact attgctttga
1381 tcttatcttg atgctcatag ctataatgaa atgccaaaca agcaccccaa tcatggccga
1441 caaaaatgat cttctttggt aaattaagaa gttcaaacca tgcagtaaga tatttgtaat
1501 gatcaagtaa cctataagaa ccattaccag atttgcctga tttgcccata ccaataaggt
1561 ctggtataat acaccgcgct actggctcaa tatgtggcac aacatgtcgc cataaataag
1621 aagaggccgc gttaccatgt aaaaaaataa cagcattttc tgcatgtttt tctgaatcat
1681 aataattaat aaatgaatca agaacattca tttgtttaca tctggcccac cactgcggac
1741 cagttatcat ccgtttcctt tgttctggat cataaacttt cgaagtcatg tcgacctcga
1801 gttgagagtg aatatgagac tctaattgga taccgagggg aatttatgga acgtcagtgg
1861 agcatttttg acaagaaata tttgctagct gatagtgacc ttaggcgact tttgaacgcg
1921 caataatggt ttctgacgta tgtgcttagc tcattaaact ccagaaaccc gcggctcagt
1981 ggctccttca acgttgcggt tctgtcagtt ccaaacgtaa aacggcttgt cccgcgtcat
2041 cggcgggggt cataacgtga ctcccttaat tctccgctca tgatcgataa cattaacgct
2101 tacaatttcc attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc
2161 tcttcgctat tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta
2221 acgccagggt tttcccagtc acgacgttgt aaaacgacgg ccagtgagcg cgcgtaatac
2281 gactcactat agggcgaatt gggtaccggg cccacccccc tactccaaaa atgtcaaaga
2341 tacagtctca gaagaccaaa gggctattga gacttttcaa caaagggtaa tttcgggaaa
2401 cctcctcgga ttccattgcc cagctatctg tcacttcatc gaaaggacag tagaaaagga
2461 aggtggctcc tacaaatgcc atcattgcga taaaggaaag gctatcattc aagatgcctc
2521 tgccgacagt ggtcccaaag atggaccccc acccacgagg agcatcgtgg aaaaagaaga
2581 cgttccaacc acgtcttcaa agcaagtgga ttgatgtgac atctccactg acgtaaggga
2641 tgacgcacaa tcccactatc cttcgcaaga cccttcctct atataaggaa gtcatttcat
2701 ttggagagga cctcgagatg gaagacgcca aaaacataaa gaaaggcccg gcgccattct
2761 atccgctgga agatggaacc gctggagagc aactgcataa ggctatgaag agatacgccc
2821 tggttcctgg aacaattgct tttacagatg cacatatcga ggtggacatc acttacgctg
2881 agtacttcga aatgtccgtt cggttggcag aagctatgaa acgatatggg ctgaatacaa
2941 atcacagaat cgtcgtatgc agtgaaaact ctcttcaatt ctttatgccg gtgttgggcg
3001 cgttatttat cggagttgca gttgcgcccg cgaacgacat ttataatgaa cgtgaattgc
3061 tcaacagtat gggcatttcg cagcctaccg tggtgttcgt ttccaaaaag gggttgcaaa
3121 aaattttgaa cgtgcaaaaa aagctcccaa tcatccaaaa aattattatc atggattcta
3181 aaacggatta ccagggattt cagtcgatgt acacgttcgt cacatctcat ctacctcccg
3241 gttttaatga atacgatttt gtgccagagt ccttcgatag ggacaagaca attgcactga
3301 tcatgaactc ctctggatct actggtctgc ctaaaggtgt cgctctgcct catagaactg
3361 cctgcgtgag attctcgcat gccagagatc ctatttttgg caatcaaatc attccggata
3421 ctgcgatttt aagtgttgtt ccattccatc acggttttgg aatgtttact acactcggat
3481 atttgatatg tggatttcga gtcgtcttaa tgtatagatt tgaagaagag ctgtttctga
3541 ggagccttca ggattacaag attcaaagtg cgctgctggt gccaacccta ttctccttct
3601 tcgccaaaag cactctgatt gacaaatacg atttatctaa tttacacgaa attgcttctg
3661 gtggcgctcc cctctctaag gaagtcgggg aagcggttgc caagaggttc catctgccag
3721 gtatcaggca aggatatggg ctcactgaga ctacatcagc tattctgatt acacccgagg
3781 gggatgataa accgggcgcg gtcggtaaag ttgttccatt ttttgaagcg aaggttgtgg
3841 atctggatac cgggaaaacg ctgggcgtta atcaaagagg cgaactgtgt gtgagaggtc
3901 ctatgattat gtccggttat gtaaacaatc cggaagcgac caacgccttg attgacaagg
3961 atggatggct acattctgga gacatagctt actgggacga agacgaacac ttcttcatcg
4021 ttgaccgcct gaagtctctg attaagtaca aaggctatca ggtggctccc gctgaattgg
4081 aatccatctt gctccaacac cccaacatct tcgacgcagg tgtcgcaggt cttcccgacg
4141 atgacgccgg tgaacttccc gccgccgttg ttgttttgga gcacggaaag acgatgacgg
4201 aaaaagagat cgtggattac gtcgccagtc aagtaacaac cgcgaaaaag ttgcgcggag
4261 gagttgtgtt tgtggacgaa gtaccgaaag gtcttaccgg aaaactcgac gcaagaaaaa
4321 tcagagagat cctcataaag gccaagaagg gcggaaagat cgccgtggaa ttcctgcagt
4381 cgccatgcgg tcattacact atccctagga gaggctggcc agaattcttc aataccaccg
4441 gttttactat cttttactct tgaactagtt aatgtgtgag tagtttcccg ataagggaaa
4501 ttagggttct tatagggttt cgctcatgtg ttgagcatat aagaaaccct tagtatgtat
4561 ttgtatttgt aaaatacttc tatcaataaa atttctaatt cctaaaacca aaatccagta
4621 ctaaaatcca gatcgcggcc gccaccgcgg tggagctcca gcttttgttc cctttagtga
4681 gggttaattc cgagcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat
4741 ccgctcacaa ttccacacaa catacgagcc ggaaghcata aagtgtaaag cctggggtgc
4801 ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt tccagtcggg
4861 aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg
4921 tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg
4981 gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa
5041 cgcaggaaag aacatgaagg ccttgacagg atatattggc gggtaaacta agtcgctgta
5101 tgtgtttgtt tgagatctca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa
5161 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg
5221 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc
5281 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc
5341 ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc
5401 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg
5461 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc
5521 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga
5581 gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc
5641 tctgctgaag ccagttacct tcggaagaag agttggtagc tcttgatccg gcaaacaaac
5701 caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg
5761 atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc
5821 acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa
5881 ttaaaaatga agttttaaat caatctaaag tatatatgtg taacattggt ctagtgatta
5941 gaaaaactca tcgagcatca aatgaaactg caatttattc atatcaggat tatcaatacc
6001 atatttttga aaaagccgtt tctgtaatga aggagaaaac tcaccgaggc agttccatag
6061 gatggcaaga tcctggtatc ggtctgcgat tccgactcgt ccaacatcaa tacaacctat
6121 taatttcccc tcgtcaaaaa taaggttatc aagtgagaaa tcaccatgag tgacgactga
6181 atccggtgag aatggcaaaa gtttatgcat ttctttccag acttgttcaa caggccagcc
6241 attacgctcg tcatcaaaat cactcgcatc aaccaaaccg ttattcattc gtgattgcgc
6301 ctgagcgaga cgaaatacgc gatcgctgtt aaaaggacaa ttacaaacag gaatcgaatg
6361 caaccggcgc aggaacactg ccagcgcatc aacaatattt tcacctgaat caggatattc
6421 ttctaatacc tggaatgctg ttttccctgg gatcgcagtg gtgagtaacc atgcatcatc
6481 aggagtacgg ataaaatgct tgatggtcgg aagaggcata aattccgtca gccagtttag
6541 tctgaccatc tcatctgtaa caacattggc aacgctacct ttgccatgtt tcagaaacaa
6601 ctctggcgca tcgggcttcc catacaatcg gtagattgtc gcacctgatt gcccgacatt
6661 atcgcgagcc catttatacc catataaatc agcatccatg ttggaattta atcgcggcct
6721 tgagcaagac gtttcccgtt gaatatggct cataacaccc cttgtattac tgtttatgta
6781 agcagacagt tttattgttc atgatgatat atttttatct tgtgcaatgt aacatcagag
6841 attttgagac acaacgtggc tttgttgaat aaatcgaact tttgctgagt tgaaggatca
6901 gatcacgcat cttcccgaca acgcagaccg ttccgtggca aagcaaaagt tcaaaatcac
6961 caactggtcc acctacaaca aagctctcat caaccgtggc tccctcactt tctggctgga
7021 tgatggggcg attcaggcga tccccatcca acagcccgcc gtcgagcggg ct
//
