Functional Characterization of a Bidirectional Plant Promoter from Cotton Leaf Curl Burewala Virus Using an Agrobacterium-Mediated Transient Assay
Abstract
:1. Introduction
2. Experimental Section
2.1. Plant Material and Bacterial Strain
2.2. Isolation of CLCuBuV Bidirectional Promoter
- (i)
- 5'- CCATGGTGACTTTGGTTTAGAGACAACAAC-3' and 5'- CTGCAGTAATTCCTAGCCCTTATTACCAG-3'
- (ii)
- 5'- CTGCAGTGACTTTGGTCAATTAGAGACAAC-3' and 5'- CCATGGTAATTCCTAGCCCTTATTACCAG-3'
2.3. Plasmid Construction
2.4. Sequence Analysis
2.5. Preparation of the Agrobacterium Suspension
2.6. Agrobacterium-Mediated Infiltration
2.7. Histochemical Detection of GUS Activity
2.8. Fluorometric Determination of GUS Activity
2.9. Statistical Analysis
3. Results
3.1. Structure and Sequence Analysis
No. | TFBs a | Function of the motif | organism | Sequence | Strand | Position |
---|---|---|---|---|---|---|
1 | Box-I | LRE b | Pisum sativum | TTTCAAA | − | 110 |
TTTCAAA | − | 286 | ||||
2 | Box-II | LRE | Solanum tubersum | TGGTAATAA | + | 433 |
3 | Box-W1 | Fungal elicitor RE | Petroselinum crispum | TTGACC | − | 7 |
4 | C-Repeat/DRE | Cold and dehydration RE | Arabidopsis thaliana | TGGCCCGAC | − | 209 |
5 | CAAT-box | Core promoter elements | many | many | +,− | many |
6 | CGCG-box | Signal RE | Arabidopsis thaliana | ACGCGC | + | 226 |
GCGCGG | − | 173 | ||||
7 | CGTCA-motif | MeJA RE | Hordeum vulgare | CGTCA | − | 223 |
8 | Circadian | Circadian control | Lycopersicon esculentum | CAANNNNATC | − | 178 |
9 | ERE | Ethylene RE | Dianthus caryophyllus | ATTTCAAA | + | 110 |
10 | GATA-motif | LRE | Arabidopsis thaliana | GATA | + | many |
11 | G-box | LRE | Triticum aestivum | TCCACATGGCA | + | 74 |
12 | GC-motif | unknown | Oryza sativa | GCCGCGCCG | + | 171 |
13 | GT-1motif | LRE | Avena sativa | GGTAAT | − | 161 |
Oryza sativa | GAAAAA | + | 290 | |||
Pisum sativum | GGTAAT | + | 435 | |||
14 | HSE | HSE c | Brassica oleracea | AGAAAACTT | + | 380 |
15 | I-box | LRE | many | many | +,− | many |
16 | MBS | MYB drought RE | Arabidopsis thaliana | CAACTG | + | 20 |
TAACTG | − | 355 | ||||
TAACTG | − | 366 | ||||
17 | Skn-1_motif | Endosperm expression RE | Oryza sativa | GTCAT | − | 222 |
18 | TATA-box | Core elements located at −30 of TSS | many | many | +,− | many |
19 | TGACG-motif | MeJA RE | Horedeum volgare | TGACG | + | 223 |
20 | W-box | LRE | Arabidopsis thaliana | TTGACC | − | 7 |
TFBs | PLACE ID | PLACE accession ID | Sequence | Copy number | Description |
---|---|---|---|---|---|
Cytokinin related | ARFAT(Aux RE) | S000270 | NGATT | 2 | “ARR1-binding element” found in rice andArabidopsis;ARR1 is response regulator;N = G/A/C/T [40] |
ARR1AT | S000454 | TGTCTC | 5 | “ARF biding site” found in the promoters of primary/early response gene [41] | |
Auxin related | SURECOREATSULTR11 | S000499 | GAGAC | 8 | Core of sulfur-responsive element SURE); containing ARF binding sequence GAGACA(complementary AuxRE TGTCTC)[42] |
CATATGGMSAUR | S000370 | CATATG | 2 | Multiple auxin response modules in the soybean SAUR15A promoter [43] | |
Mesophyll-specific | CACTFTPPCA1 | S000449 | YACT | 8 | Mesophyll-specific gene expression in C4 plant Flaveria trinervia [44]. |
Pollen-specific | POLLEN1LELAT52 | S000245 | AGAAA | 1 | Pollen-specific expression of tomato Late52 gene [45] |
Root-specific | ROOTMOTIFTAPOX1 | S000098 | ATATT | 5 | Root-specific motifs found in the rolD promoter [46]. |
OSE2ROOTNODULE | S000468 | CTCTT | 1 | Nodule specificity of soybean lbc3 and N23 gene promoters [47]. |
3.2. Transient Expression of the Bidirectional Promoter
3.3. Nucleotide Sequence Comparison of the CLCuBuV LIR
No | Virus Acronym | % identity a | LIR size (bp) | Accession IDs |
---|---|---|---|---|
1 | CLCuBuV | 100 | 455 | FR837932 |
2 | CLCuMuV | 81.8 | 440 | AJ496287 |
3 | CLCuKoV | 81.6 | 447 | AJ496286 |
4 | CLCuShV | 81.9 | 447 | FN552004 |
5 | HYVMV | 77.4 | 437 | FR772082 |
6 | CLCuBaV | 76.4 | 444 | NC_007290 |
7 | MaYVCMV | 73.6 | 440 | FR715681 |
8 | PaLCuV | 70.1 | 478 | FM955602 |
9 | AEV | 69.9 | 445 | AM698011 |
10 | CYVMV | 69.8 | 451 | FN645926 |
11 | ToLCPKV | 68.1 | 456 | AM948961 |
12 | ChiLCMuV | 68.7 | 450 | FM149613 |
13 | SiLCV | 66.9 | 447 | DQ641706 |
14 | CLCuAaV | 65.2 | 434 | AJ002452 |
15 | CLCuRaV | 64.6 | 444 | JF502364 |
16 | CLCuGeV-PK | 32.7 | 453 | FR751142 |
17 | CLCuGeV-SD | 32.6 | 451 | AY036007 |
3.4. Enriched Regulatory Elements in the Geminivirus Bidirectional Promoters
3.5. Identification of TFBs that Bind to the Promoters of ds-DNA Animal Viruses
4. Discussion
5. Conclusions
Acknowledgments
Conflicts of Interest
References and Notes
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Ashraf, M.A.; Shahid, A.A.; Rao, A.Q.; Bajwa, K.S.; Husnain, T. Functional Characterization of a Bidirectional Plant Promoter from Cotton Leaf Curl Burewala Virus Using an Agrobacterium-Mediated Transient Assay. Viruses 2014, 6, 223-242. https://doi.org/10.3390/v6010223
Ashraf MA, Shahid AA, Rao AQ, Bajwa KS, Husnain T. Functional Characterization of a Bidirectional Plant Promoter from Cotton Leaf Curl Burewala Virus Using an Agrobacterium-Mediated Transient Assay. Viruses. 2014; 6(1):223-242. https://doi.org/10.3390/v6010223
Chicago/Turabian StyleAshraf, Muhammad Aleem, Ahmad Ali Shahid, Abdul Qayyum Rao, Kamran Shehzad Bajwa, and Tayyab Husnain. 2014. "Functional Characterization of a Bidirectional Plant Promoter from Cotton Leaf Curl Burewala Virus Using an Agrobacterium-Mediated Transient Assay" Viruses 6, no. 1: 223-242. https://doi.org/10.3390/v6010223