Open Access Highly Accessed Research article

Larval midgut modifications associated with Bti resistance in the yellow fever mosquito using proteomic and transcriptomic approaches

Guillaume Tetreau1*, Krishnareddy Bayyareddy2, Christopher M Jones3, Renaud Stalinski1, Muhammad A Riaz1, Margot Paris1, Jean-Philippe David1, Michael J Adang24 and Laurence Després1

Author Affiliations

1 Laboratoire d’Ecologie Alpine, LECA-UMR 5553, Université de Grenoble 1, BP 53, 38041, Grenoble cedex 09, France

2 Department of Entomology, University of Georgia, Athens, GA, 30602-2603, USA

3 Vector Group, Liverpool School of Tropical Medicine, Liverpool, L3 5QA, UK

4 Department of Biochemistry and Molecular Biology, University of Georgia, Athens, GA, 30602-2603, USA

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BMC Genomics 2012, 13:248  doi:10.1186/1471-2164-13-248

Published: 15 June 2012

Additional files

Additional file 1:

All the 3512 transcripts detected by microarrays experiments in at least 5 hybridizations out of 6. For each transcript, accession number, corrected p-value, expression level changes, Vectorbase annotation and functional category are indicated.

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Additional file 2:

70 transcripts significantly (corrected P-val<0.01) more than 3-fold differentially transcribed in the LiTOX strain. Transcripts are classified according to their putative function using the 13 functional categories. For each transcript, accession number, corrected P-value, expression level changes, Vectorbase annotation and supercontig are indicated. For transcripts of ‘unknown functions’, their putative function with corresponding score, ID, accession number and species of the best hit found using BLASTP software are indicated.

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Additional file 3:

Validation of microarray data by RT-qPCR on fifteen selected genes. Both experiments were performed on the same mRNA extracted from dissected larval midguts. ALP2, Alkaline phosphatase AAEL003298; ALP3, AAEL003313; ALP5, AAEL015070; ALP6, AAEL011175; APN1, N-Aminopeptidase AAEL012774; APN2, AAEL012776; APN3, AAEL012778; Cad2, Cadherin AAEL007488; HP1, Conserved hypothetical protein AAEL010435; HP2, AAEL013584; SE1, Serine-type endopeptidase AAEL007938; SE2, Serine-type endopeptidase AAEL011917; Cytochrome P450: CYP6Z7, AAEL009130; CYP6Z8, AAEL009131 and CYP4D24, AAEL007815.

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Additional file 4:

2D-DIGE gels from the two biological replicates and dye-swapping. BBMV prepared from first (A and B) and second (C and D) biological replicate are separated in function of their size (kDa) and their isoelectric point (pI). BBMV from Bti resistant strain are labeled with Cy3 and susceptible strain with Cy5 (A and C) or resistant strain with Cy5 and susceptible with Cy3 (B and D).

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Additional file 5:

Protein identification of the 56 spots picked on deep purple stained 2D-gel. When different spots pointed to the same protein, they were differentiated using arbitrary letters after the access number. For each identification, the predicted pI, the predicted mass in kilodaltons, the percentage of sequence coverage, their functional category, and the species and database matched are indicated.

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Additional file 6:

Glycosylphosphatidylinositol (GPI)-anchor domains detection by four predictive computational programs. For each gene and protein, their accession number, the transcript and protein sizes are indicated. Results from the big-GPI and GPI-SOM softwares are indicated as ‘YES’ when they found a potential GPI-domain and ‘NO’ when no GPI-domain was determined. For PredGPI, presence is indicated by ‘Highly probable’, ‘Weakly probable’ or ‘Probable’ and absence by ‘NO’. For FragAnchor, presence of GPI domain is indicated by ‘Highly probable’ or ‘Probable’, absence by ‘NO’ and when prediction is uncertain by ‘Potential false positive’.

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Additional file 7:

Cadherin detection by immunoblotting. BBMV proteins from the susceptible Bora-Bora strain (lane 1), LiTOX strain (lane 2) and the UGAL Aedes strain (lane 3) were separated in SDS-PAGE and stained with coomassie blue (panel A) or probed with α-AgCad1 antibodies (panel B), α-AgCad2 antibodies (panel C) or with pre-immune serum from α-AgCad2 rabbit (panel D).

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Additional file 8:

Primer pairs used for RT-qPCR analyses. For each primer pair, sequence, corresponding gene name and accession number, product length, Tm and optimal annealing temperature used in PCR program are indicated. PCR efficiency and different parameters of the calibration curves (R2, slope and y-intercept) are also indicated. Specificity of each primer pair was first assessed by BLAST analysis against Ae. aegypti genome and then verified by performing a melt curve analysis. A high specificity is indicated as “YES” when the primer pair matched to a unique position in the Ae. aegypti genome and when PCR product Tm was correct.

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