Monitorització de ruscs d'abelles

L’apicultura és una de les especialitzacions de la ramaderia que a l’actualitat encara es manté sense gairebé cap tipus de tecnologia associada. Els ruscs continuen funcionant de forma similar a fa milers d’anys, on les abelles fan la feina i l’apicultor recol·lecta els productes que en generen. El present treball consisteix en dissenyar i construir un prototip que permeti monitoritzar diferents paràmetres d’un rusc d’abelles: la temperatura interna i externa, la humitat també interna i externa així com comptabilitzar el nombre d’abelles que entren i surten del rusc. Totes les dades que es van adquirint queden emmagatzemades en una memòria SD. Per aconseguir-ho s’ha dissenyat i creat un prototip, de baix cost i de fàcil implementació e instal·lació. En aquest prototip en qualsevol moment l’usuari pot observar els diferents paràmetres monitoritzats mitjançant una pantalla LCD. Per a comprovar el correcte funcionament del prototipus, s’han realitzat proves de camp, incorporant-lo a un rusc d’abelles reals i prenent mesures durant un temps limitat. El treball s’ha realitzat seguint una planificació prèviament definida, començant per un estudi a fons del mercat, realitzant diverses consultes a experts en l’apicultura, el disseny i muntatge del prototip, la seva programació i finalment l’anàlisi dels resultats obtinguts a les proves de camp. Els resultats mostren la viabilitat del prototipus creat i dona peu a la seva utilització per part d’apicultors interessats en conèixer quelcom més de l’estat del rusc o per a la utilització a l’estudi del comportament de les abelles

Additional file 2: Figure S1. of Monocot and dicot MLO powdery mildew susceptibility factors are functionally conserved in spite of the evolution of class-specific molecular features

Expression levels of PsMLO1 and HvMLO after transformation. Panel A) and panel B) show the expression of PsMLO1 and HvMLO in 19 and 20 T1 individuals, respectively, which were obtained by the transformation of the tomato mutant line Slmlo1, harboring a loss-of-function mutation of the endogenous SlMLO1 gene. Asterisks indicate T1 individuals selected for self-pollination and the development of T2 families. (PDF 173 kb

Additional file 3: Figure S2. of Monocot and dicot MLO powdery mildew susceptibility factors are functionally conserved in spite of the evolution of class-specific molecular features

Effects of transgenic overexpression of pea PsMLO1 and barley HvMLO in the tomato mutant line Slmlo1. Average disease index (DI) values and phenotypes are referred to transgenic plants of two additional T2 families segregating for PsMLO1 [35S::PsMLO1-4 and 35S::PsMLO1-6, panel a) and b)] and two additional T2 families segregating for HvMLO [35S::HvMLO-10 and 35S::HvMLO-15), panel c) and d)]. Data relative to the Slmlo1 mutant line, used as genetic background for transformation, and non-transgenic plants of three T2 families for each overexpression construct (35S::PsMLO1_(−) and 35S::HvMLO_(−)) are also shown. (PDF 304 kb

Assessment of <i>Leveillula taurica</i> susceptibility of <i>ol-2::CaMlo2</i> transformants.

<p><b>A.</b><i>L. taurica</i> disease symptoms on tomato mutant <i>ol-2</i> and two individual T2 plants of <i>CaMlo2</i>-overexpressing transformants of <i>ol-2</i>. <b>B.</b> and <b>C.</b> <i>L. taurica</i> biomass and <i>CaMlo2</i> expression level quantified by qRT-PCR of T2 plants expressing <i>CaMlo2</i> compared with <i>ol-2</i> plants for T2 family 1 (<b>B</b>) and T2 family 6 (<b>C</b>). Columns refer to relative quantification with respect to the <i>ol-2</i> plants. Bars refer to standard errors of the mean of 11 T2 plants of family 1, 12 T2 plants of family 6, and 10 <i>ol-2</i> plants. The ΔΔCt method with <i>SlEf</i> as the reference gene was used for normalization. DNA and RNA were isolated from pooled 2^nd and 3^rd leaves of each plant. Asterisks refer to significant differences with respect to <i>ol-2</i> plants, inferred by means comparison by Student's t-test.</p

Effects of virus induced gene silencing (VIGS) for pepper <i>CaMlo1</i> and <i>CaMlo2</i>.

<p><b>A.</b> Quantification of <i>CaMlo1</i> and <i>CaMlo2</i> gene expression in pepper plants (cv. A) subjected to VIGS. Two silencing constructs for each gene were developed (VIGS:CaMlo1-a and -b; VIGS:CaMlo2-a and -b). Columns refer to transcripts fold change with respect to plants inoculated with an empty TRV2 vector (EV plants). RNAs were isolated from pooled tissues of the 4^th, 5^th and 6^th leaf for each plant. Relative quantification was performed by using the ΔΔCt method and the reference gene <i>CaActin</i>. Bars refer to standard errors of the mean of three biological replicates. Asterisks refer to significant differences with respect to expression levels of EV plants, inferred by means comparison with a Student's t-test. <b>B</b>. Quantification of <i>L. taurica</i> colonization levels on EV, VIGS:CaMlo1 (a and b) and VIGS:CaMlo2 (a and b) pepper plants (cv. A) by counting the average colony number on the 4^th, 5^th and 6^th leaf for each plant. Bars refer to the standard error of the mean of seven plants for each treatment. Asterisks refer to significant differences with respect to EV plants, inferred by means comparison by Student's t-test. <b>C</b>. Quantification of <i>L. taurica</i> colonization levels on EV, VIGS:CaMlo1 (a and b) and VIGS:CaMlo2 (a and b) pepper plants (cv. A) by of fungal biomass by real time qPCR. DNAs were isolated from pooled 4^th, 5^th and 6^th infected leaves per plant. In total, seven plants were tested for each VIGS vector. The ΔΔCt method with <i>CaActin</i> as the reference gene was used for normalization. Columns refer to relative quantification with respect to EV plants. Bars refer to standard errors referred to seven biological replicates. Asterisks refer to significant differences with respect to EV plants, inferred by means comparison by Student's t-test.</p

Multiple amino acid sequence alignment of pepper CaMLO1 and CaMLO2, barley HvMLO, tomato SlMLO1, as well as <i>Arabidopsis</i> AtMLO2, AtMLO6, and AtMLO12.

<p>The alignment was generated by CLUSTALW using default parameters. Grey boxes indicate similar amino acids and black boxes indicate identical amino acids. The C-terminal D/E-F-S/T-F tetra-peptide sequence, one of several motifs characteristic of barley <i>Mlo</i> orthologs <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0070723#pone.0070723-Panstruga1" target="_blank">[30]</a> is outlined in grey. For this tetra-peptide sequence, CaMLO2 contains D-F-T-F, which is identical to SlMLO1.</p

Results from artificial inoculation of <i>Leveillula taurica</i> on different tomato genotypes.

<p><b>A.</b> From left to right: transformant of the breeding line <i>ol-2</i> in which the wild-type <i>SlMlo1</i> cDNA is overexpressed (<i>ol-2</i>::<i>SlMlo1</i>); cultivar Moneymaker (MM), homozygous for the wild-type <i>SlMlo1</i> allele; breeding line <i>ol-2,</i> homozygous for a <i>Slmlo1</i> loss-of-function allele; cultivar Laurica, carrying the <i>Lv</i> gene for hypersensitive response-based resistance. Yellow patches on the adaxial surface of <i>ol-2::SlMlo1</i>, MM and <i>ol-2</i> leaves are due to <i>L. taurica</i> colonization and correspond to abaxial fungal sporulation. Necrotic lesions on the Laurica genotype are the consequence of Lv-gene-mediated hypersensitive response and do not correspond to fungal growth on the abaxial side of the leaves. Pictures were taken four weeks after fungal inoculation. The experiment was carried out twice yielding similar results. <b>B.</b> Fungal DNA quantification by real-time PCR on the same genotypes described above. Amount of <i>L. taurica</i> DNA was normalized by the plant reference gene elongation factor (<i>SlEf</i>) with the ΔΔCt method. The relative pathogen biomass in MM plants is set as 1. In total, three leaves (4^th, 5^th and 6^th leaf) per plant were pooled together, and three plants per genotype were assayed. Bars indicate the standard error of the mean for three biological replicates. Asterisks indicate significant difference with the control cultivar MM, performed by Student's t-test.</p

Accumulation of <i>CaMlo1</i> and <i>CaMlo2</i> transcripts during <i>Leveillula taurica</i> infection in pepper.

<p><b>A.</b> Expression profile of <i>CaMlo</i> genes measured by real time qRT-PCR in pepper (cultivar A) leaves upon <i>Leveillula taurica</i> infection. Columns indicate transcript fold changes with respect to non-inoculated plants (0 hours after inoculation, hpi). Relative quantification was performed by using the ΔΔCt method and the reference gene <i>CaUEP</i>. Samples were taken from three whole pepper leaves per plant (the 3^rd, 4^th and 5^th leaf) upon <i>L. taurica</i> infection at the following time points: 0 hpi, 1 hpi, 3 hpi, 5 hpi, 7 hpi, 21 hpi, 25 hpi, 30 hpi, 47 hpi, 72 hpi, 96 hpi and 21 days post inoculation (dpi). Results are based on three individual pepper plants per time point. Bars refer to standard errors of the biological replicates and asterisks refer to significant differences with respect to non-inoculated plants (0 hpi), inferred by mean comparisons with a Student's t-test. Time spans named as B1, B2, B3 and B4 refer to the fungal infection stages as described in the text and in panel B. Expression analyses were carried out with a different primer set for each <i>CaMlo</i> gene. <b>B.</b> Infection process of the endophytic powdery mildew <i>Leveillula taurica</i> on surface of pepper leaves. <b>1</b>, A conidium (c) germinates and a primary adhesion body (pab) is formed at the tip of the germ tube. <b>2</b>, Primary (infection) hyphae (ih) grow into the stomata (s) and secondary adhesion bodies (sab) are formed on the secondary hyphae (sh). <b>3</b>, The pathogen grows in the intercellular space and haustoria (h) are formed in mesophyll cells. <b>4</b>, Conidiophores (cp) are projected from the stomata three weeks after inoculation and superficial hyphae (sh) elongate on both sides of the leaves for a new round of infection.</p

Expression of <i>CaMlo1</i> and <i>CaMlo2</i> in pepper upon <i>Leveillula taurica</i> infection measured by semi-quantitative RT-PCR at five time points: 0-hpi (hours post inoculation), 5 hpi, 25 hpi, 72 hpi and 21 dpi (days post inoculation).

<p>Leaf samples were collected from (in a left to right order): 1, susceptible cultivar A; 2, susceptible cultivar B; 3, susceptible cultivar Maor; 4, resistant doubled haploid HV-12; 5, Maor; 6, cultivar A; 7, HV-12; 8, cultivar B. Samples from different time points were separated by 1Kb marker (M). In total, three leaves per plant from each genotype were collected and leaves from each plant were pooled for RNA isolation. The analysis was carried out with a second primer set for <i>CaMlo1</i> and <i>CaMlo2</i>, yielding similar trends for expression.</p

DataSheet_1_ZED1-related kinase 13 is required for resistance against Pseudoidium neolycopersici in Arabidopsis accession Bla-6.docx

To explore specific components of resistance against the tomato-adapted powdery mildew pathogen Pseudoidium neolycopersici (On) in the model plant Arabidopsis, we performed a disease assay in 123 accessions. When testing the resistance in the F1 from crossings between resistant accessions with susceptible Col-0 or Sha, only the progeny of the cross between accession Bla-6 and Col-0 displayed a completely resistant phenotype. The resistance in Bla-6 is known to be specific for Pseudoidium neolycopersici. QTL analysis and fine-mapping through several rounds of recombinant screenings allowed us to locate a major resistance QTL in an interval on chromosome 1, containing two candidate genes and an intergenic insertion. Via CRISPR/Cas9 targeted mutagenesis, we could show that knocking out the ZED-1 RELATED KINASE 13 (ZRK13) gene compromised the On resistance in Bla-6. Several polymorphisms are observed in the ZRK13 allelic variant of Bla-6 when compared to the Col-0 protein.</p