| 1. |
- Beal, Jacob, et al.
(författare)
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Robust estimation of bacterial cell count from optical density
- 2020
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Ingår i: Communications Biology. - : Springer Science and Business Media LLC. - 2399-3642. ; 3:1
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Tidskriftsartikel (refereegranskat)abstract
- Optical density (OD) is widely used to estimate the density of cells in liquid culture, but cannot be compared between instruments without a standardized calibration protocol and is challenging to relate to actual cell count. We address this with an interlaboratory study comparing three simple, low-cost, and highly accessible OD calibration protocols across 244 laboratories, applied to eight strains of constitutive GFP-expressing E. coli. Based on our results, we recommend calibrating OD to estimated cell count using serial dilution of silica microspheres, which produces highly precise calibration (95.5% of residuals <1.2-fold), is easily assessed for quality control, also assesses instrument effective linear range, and can be combined with fluorescence calibration to obtain units of Molecules of Equivalent Fluorescein (MEFL) per cell, allowing direct comparison and data fusion with flow cytometry measurements: in our study, fluorescence per cell measurements showed only a 1.07-fold mean difference between plate reader and flow cytometry data.
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| 2. |
- Saevarsdottir, S., et al.
(författare)
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Multiomics analysis of rheumatoid arthritis yields sequence variants that have large effects on risk of the seropositive subset
- 2022
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Ingår i: Annals of the Rheumatic Diseases. - : BMJ. - 0003-4967 .- 1468-2060. ; 81:8
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Tidskriftsartikel (refereegranskat)abstract
- Objectives To find causal genes for rheumatoid arthritis (RA) and its seropositive (RF and/or ACPA positive) and seronegative subsets. Methods We performed a genome-wide association study (GWAS) of 31 313 RA cases (68% seropositive) and similar to 1 million controls from Northwestern Europe. We searched for causal genes outside the HLA-locus through effect on coding, mRNA expression in several tissues and/or levels of plasma proteins (SomaScan) and did network analysis (Qiagen). Results We found 25 sequence variants for RA overall, 33 for seropositive and 2 for seronegative RA, altogether 37 sequence variants at 34 non-HLA loci, of which 15 are novel. Genomic, transcriptomic and proteomic analysis of these yielded 25 causal genes in seropositive RA and additional two overall. Most encode proteins in the network of interferon-alpha/beta and IL-12/23 that signal through the JAK/STAT-pathway. Highlighting those with largest effect on seropositive RA, a rare missense variant in STAT4 (rs140675301-A) that is independent of reported non-coding STAT4-variants, increases the risk of seropositive RA 2.27-fold (p=2.1x10(-9)), more than the rs2476601-A missense variant in PTPN22 (OR=1.59, p=1.3x10(-160)). STAT4 rs140675301-A replaces hydrophilic glutamic acid with hydrophobic valine (Glu128Val) in a conserved, surface-exposed loop. A stop-mutation (rs76428106-C) in FLT3 increases seropositive RA risk (OR=1.35, p=6.6x10(-11)). Independent missense variants in TYK2 (rs34536443-C, rs12720356-C, rs35018800-A, latter two novel) associate with decreased risk of seropositive RA (ORs=0.63-0.87, p=10(-9)-10(-27)) and decreased plasma levels of interferon-alpha/beta receptor 1 that signals through TYK2/JAK1/STAT4. Conclusion Sequence variants pointing to causal genes in the JAK/STAT pathway have largest effect on seropositive RA, while associations with seronegative RA remain scarce.
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| 3. |
- Hamada, Afaf M., et al.
(författare)
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Thiamine treatments alleviate aphid infestations in barley and pea
- 2013
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Ingår i: Phytochemistry. - : Elsevier BV. - 0031-9422 .- 1873-3700. ; 94, s. 135-141
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Tidskriftsartikel (refereegranskat)abstract
- Treatment of plants with thiamine (Vitamin B1) has before been shown to activate plant defence against microorganisms. Here, we have studied the effects of thiamine treatments of plants on aphid reproduction and behaviour. The work was mainly carried out with bird cherry-oat aphid (Rhopalosiphum padi L.) on barley (Hordeum vulgare L.). Aphid population growth and aphid acceptance on plants grown from seeds soaked in a 150 mu M thiamine solution were reduced to ca. 60% of that on control plants. R. padi life span and the total number of offspring were reduced on barley plants treated with thiamine. Healthy aphids and aphids infected with the R. padi virus were similarly affected. Spraying or addition of thiamine at 150 mu M to nutrient solutions likewise resulted in reduced aphid population growth to ca. 60%, as did plant exposure to thiamine odour at 4 mM. Thiamine treatments resulted in reduced aphid population growth also when tested with grain aphid (Sitobion avenae F.) on barley and pea aphid (Acyrthosiphon pisum H.) on pea (Pisum sativum L). There was no direct effect of thiamine on aphid reproduction or thiamine odour on aphid behaviour, as evaluated using artificial diets and by olfactometer tests, respectively. Two gene sequences regulated by salicylic acid showed higher transcript abundance and one gene sequence regulated by methyl jasmonate showed lower transcript abundance in thiamine-treated plants but not in control plants after aphid infestation. These results suggest that the aphid antibiosis and antixenosis effects may be related to priming of defence, but more studies are needed to explain the effects against aphids.
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| 4. |
- Delp, Gabriele, et al.
(författare)
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Microarray analysis of the interaction between the aphid Rhopalosiphum padi and host plants reveals both differences and similarities between susceptible and partially resistant barley lines
- 2009
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Ingår i: Molecular Genetics and Genomics. - : Springer Science and Business Media LLC. - 1617-4615 .- 1617-4623. ; 281:3, s. 233-248
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Tidskriftsartikel (refereegranskat)abstract
- The bird cherry-oat aphid (Rhopalosiphum padi L.) is an important pest on cereals causing plant growth reduction without specific leaf symptoms. Breeding of barley (Hordeum vulgare L.) for R. padi resistance shows that there are several resistance genes, reducing aphid growth. To identify candidate sequences for resistance-related genes, we performed microarray analysis of gene expression after aphid infestation in two susceptible and two partially resistant barley genotypes. One of the four lines is a descendant of two of the other genotypes. There were large differences in gene induction between the four lines, indicating substantial variation in response even between closely related genotypes. Genes induced in aphid-infested tissue were mainly related to defence, primary metabolism and signalling. Only 24 genes were induced in all lines, none of them related to oxidative stress or secondary metabolism. Few genes were down-regulated, with none being common to all four lines. There were differences in aphid-induced gene regulation between resistant and susceptible lines. Results from control plants without aphids also revealed differences in constitutive gene expression between the two types of lines. Candidate sequences for induced and constitutive resistance factors have been identified, among them a proteinase inhibitor, a serine/threonine kinase and several thionins.
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| 5. |
- Larsson, Kristina A. E., et al.
(författare)
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Differential regulation of 3-aminomethylindole/N-methyl-3-aminomethylindole N-methyltransferase and gramine in barley by both biotic and abiotic stress conditions
- 2011
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Ingår i: Plant physiology and biochemistry (Paris). - : Elsevier BV. - 0981-9428 .- 1873-2690. ; 49:1, s. 96-102
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Tidskriftsartikel (refereegranskat)abstract
- The expression of NMT (3-aminomethylindole/N-methyl-3-aminomethylindole N-methyltransferase; EC 2.1.1.), involved in the biosynthesis of the indole alkaloid gramine, was investigated in aphid-infested barley (Hordeum vulgare L). NMT is induced by methyl jasmonate and it was hypothesized that the gene would be more strongly upregulated in aphid-resistant barley. We examined the effects of feeding by three aphid species; Russian wheat aphid (Diuraphis noxia Mordvilko), rose-grain aphid (Metopolophium dirhodum Walker) and bird cherry-oat aphid (Rhopalosiphum padi L.) on barley genotypes with varying resistance characteristics. The barley genotypes selected included the cultivar Libra, known to upregulate gramine after feeding by Schizaphis graminum. Infestation by R. padi and M. dirhodum resulted in higher NMT expression in the doubled haploid line 5172-28:4 (DH28:4), which has moderate resistance against R. padi, but not in other aphid barley combinations. None of the aphid plant combinations had however increased gramine, suggesting that aphid-induction of gramine is specific to S. graminum. The increased abundance of NMT transcript in aphid-infested DH28:4 did not lead to higher amounts of NMT protein or NMT enzyme activity, neither did 200 times upregulation of NMT transcript in cotyledons incubated with methyl jasmonate, illustrating that even large differences measured at transcript level may have no metabolic consequences. Drought stress or treatments with abscisic acid did lead to higher gramine concentrations in several barley cultivars, but without any concomitant increase of NMT transcripts. Thus, the regulation of the biosynthetic pathway to gramine at transcript and metabolite level diverges during two different stress conditions.
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| 6. |
- Larsson, Kristina A. E., et al.
(författare)
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N-Methyltransferase involved in gramine biosynthesis in barley : Cloning and characterization
- 2006
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Ingår i: Phytochemistry. - : Elsevier BV. - 0031-9422 .- 1873-3700. ; 67:18, s. 2002-2008
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Tidskriftsartikel (refereegranskat)abstract
- The indole alkaloid gramine occurs in leaves of certain barley (Hordeum vulgare L.) cultivars but not in others. A gene sequence in barley that earlier was characterized as a jasmonate-induced O-methyltransferase (MT) (EC 2.1.1.6, GenBank accession U54767) was here found to be absent in some barley cultivars and breeding lines that all lacked gramine. The cDNA was cloned and expressed in Escherichia coli and the recombinant protein purified. The purified recombinant protein methylated two substrates in the pathway to gramine: 3-aminomethylindole (AMI) and N-methyl-3-aminomethylindole (MAMI) at a high rate, with Km-values of 77 mu M and 184 mu M, respectively. In contrast, the protein did not exhibit any detectable methylation with the earlier suggested substrate for O-methylation, caffeic acid. A number of cultivars and breeding lines of barley were analyzed for presence of the U54767 gene sequence and NIT protein and the enzyme activity in vitro with MAMI or caffeic acid as substrates. The results showed a clear relationship between the presence of the MT gene, the NIT protein and N-methyltransferase activity, and confirmed the identification of the gene as coding for an N-methyltransferase (NMT, EC 2.1.1) and being involved in gramine biosynthesis.
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| 7. |
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| 8. |
- Mehrabi, Sara, 1982-, et al.
(författare)
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The constitutive expression and induction of three β-1,3-glucanases by bird cherry-oat aphid in relation to aphid resistance in 15 barley breeding lines
- 2016
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Ingår i: Arthropod-Plant Interactions. - : Springer Science and Business Media LLC. - 1872-8855 .- 1872-8847. ; 10:2, s. 101-111
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Tidskriftsartikel (refereegranskat)abstract
- In this study, 15 closely related barley genotypes were analyzed for the abundance of three β-1,3-glucanase transcripts immediately before and during infestation by the bird cherry-oat aphid (Rhopalosiphum padi L.). The barley lines are doubled haploid lines in backcross (BC) generations BC1 and BC2 from a cross between cultivar Lina and a wild barley accession. Previously, they have been characterized as susceptible (S) or resistant (R) to R. padi based on their ability to support nymphal growth. Here we also tested whether resistance was manifested as reduced aphid settling on the plants. Indeed, aphid numbers were lower on R than on S lines in all cases where there were significant differences between R and S lines. The choice of β-1,3-glucanase sequences is based on earlier results comparing two S and two R genotypes, suggesting that at least two of the three studied sequences are susceptibility factors. The comparisons of transcript abundance in plants with aphids showed for two of the β-1,3-glucanase sequences that there were several cases where an S genotype had significantly higher abundance than an R genotype, and in no case did an R line have significantly higher abundance than an S line. Thus, there was some support for the idea that β-1,3-glucanase sequences are susceptibility factors in the interaction between barley and R. padi.
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| 9. |
- Mehrabi, Sara, et al.
(författare)
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Transcript abundance of resistance- and susceptibility-related genes in a barley breeding pedigree with partial resistance to the bird cherry-oat aphid (Rhopalosiphum padi L.)
- 2014
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Ingår i: Euphytica. - : Springer Science and Business Media LLC. - 0014-2336 .- 1573-5060. ; 198:2, s. 211-222
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Tidskriftsartikel (refereegranskat)abstract
- Aphids such as the bird cherry-oat aphid (Rhopalosiphum padi L.) are serious pests in agriculture worldwide. Partially resistant barley genotypes have been produced in a breeding program for resistance to R. padi. The resistance is manifested as smaller aphids after a certain period of nymphal growth on the test plants. A large number of doubled haploid breeding lines thereby characterized as partially resistant or susceptible to R. padi are available from this breeding program. In the present study a selection of these lines, spanning four generations and 23 genotypes, was used for the evaluation of the constitutive transcript abundance of gene sequences in young barley plants. The genes were selected from an earlier microarray study where they had been identified as being up-regulated by aphids in all four barley genotypes studied or differentially up-regulated or constitutively expressed in the two resistant as compared to the two susceptible barley genotypes. The prediction for constitutive transcript abundance across the selection of genotypes was here confirmed for five out of eight investigated genes. Two of these were, as predicted, expressed at equal levels in all genotypes. Two gene sequences exhibited higher transcript abundance in resistant than in susceptible offspring; one coding for a thionin and another for a proteinase inhibitor. A lipoxygenase gene had higher transcript abundance in the susceptible lines. The study thus gives further support for putative constitutive roles of three genes previously identified as significant in the barley-R. padi interaction.
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| 10. |
- Nikus, J, et al.
(författare)
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Cloning of a plastidic rye (Secale cereale) beta-glucosidase cDNA and its expression in Escherichia coli
- 2003
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Ingår i: Physiologia Plantarum. - : Wiley. - 0031-9317 .- 1399-3054. ; 118:3, s. 337-345
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Tidskriftsartikel (refereegranskat)abstract
- The cDNA for a beta-glucosidase (EC3.2.1.21) was isolated from rye (Secale cereale, cv Motto) and the sequence corresponding to the mature protein cloned into pET21a expression vector and used for transformation of Escherichia coli. The recombinant beta-glucosidase expressed in E. coli was recognized by antibodies to maize beta-glucosidase and exhibited the same kinetic properties on the endogenous substrates hydroxamic acid glucosides and artificial substrates as the native enzyme purified from rye. The enzyme monomer had an apparent molecular weight of about 67 kDa. The isolated cDNA was analysed with web-based chloroplast targeting prediction programs. The programs predicted a chloroplast targeting peptide with a cleavage site between amino acid 49 and 50. Sequence alignment of the plastidic rye beta-glucosidase showed that the putative sites for substrate specificity of maize Glu1, W378 and F198 (F197) are conserved in the rye enzyme, whereas F205, F466 and A467 of maize Glu1 are exchanged for histidine, glycine and serine, respectively, in rye. The plastidic beta-glucosidase is expressed in all plant parts and the highest levels were found in the coleoptile and mesocotyl.
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| 11. |
- Saheed, Sefiu A., et al.
(författare)
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Russian wheat aphid causes greater reduction in phloem transport apacity of barley leaves than bird cherry-oat aphid.
- 2010
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Ingår i: Acta Botanica Croatica. - 0365-0588 .- 1847-8476. ; 69:1, s. 7-18
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Tidskriftsartikel (refereegranskat)abstract
- The effects of feeding by the Russian wheat aphid (RWA). Dutraplu.s nom! ordvilko and the Bird cherry-oat aphid (BC A). Rhopaloppluan pad, L on he transport capacity of barley Holdeum vulgare L leaves were nvestigated and compated with a view to i-elan ng these effects to the isible symptoms shown by the respective infested plants RWA causes xtensive chlorosis and neciosis on an infested plant whereas I3CA auses no obseivable symptoms Our results using the xenobiotic. phloem obile Bum ophole. 5, 6 carboxyBurn escei n chacetate (5. 6-CFDA) evealed striking ch fletences in damage to the transpol t of ssimilates thiough the phloem by these two aphids The result clearly uggests that short-term feeding by RWA causes a reduction in tiansport f assimikites and a mole severe reduction oi pei haps even permanent essation of transport during long-term feed111,2. In contrast. feechntz y BCA does not lead to a !milked dect ease in transport do ring hort-term feeding period. howevei, a !eduction in the uanspoit was ecorded donne long-term feeding activities These iesults perhaps uggest that damage to ti ansport capacities of the barley leaves ppeals lobe partly responsible for the observed symptoms in WIA-infested plants and the lack of them during BCA in symptoms such as eduction ol cessation in transport of assionlates to growing tissues ay lead to such observable symptoms
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| 12. |
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| 13. |
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| 14. |
- Saheed, Sefiu, et al.
(författare)
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Stronger induction of callose deposition in barley by Russian wheat aphid than bird cherry-oat aphid is not associated with differences in callose synthase or beta-1,3-glucanase transcript abundance
- 2009
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Ingår i: Physiologia Plantarum. - : Wiley. - 0031-9317 .- 1399-3054. ; 135:2, s. 150-161
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Tidskriftsartikel (refereegranskat)abstract
- The effects of infestation by the bird cherry-oat aphid (BCA), (Rhopalosiphum padi L) and the Russian wheat aphid (RWA) (Diuraphis noxia Mordvilko) on callose deposition and transcription of genes related to callose accumulation were investigated in barley (Hordeum vulgare L. cv. Clipper). The BCA, which gives no visible symptoms, induced very limited callose deposition, even after 14 days of infestation. In contrast, RWA, which causes chlorosis, white and yellow streaking and leaf rolling, induced callose accumulation already after 24 h in longitudinal leaf veins. The deposition was pronounced after 72 h, progressing during 7 and 14 days of infestation. In RWA-infested source leaves, callose was also induced in longitudinal veins basipetal to the aphid-infested tissue, whereas in sink leaves, more callose deposition was found above the feeding sites. Eight putative callose synthase genes were identified in a database search, of which seven were expressed in the leaves, but with similar transcript accumulation in control and aphid-infested tissue. Five out of 12 examined beta-1,3-glucanases were expressed in the leaves. All five were upregulated in RWA-infested tissue, but only two in BCA-infested tissue, and to a lesser extent than by RWA. The results suggest that callose accumulation may be partly responsible for the symptoms resulting from RWA infestation and that a callose-inducing signal may be transported in the phloem. Furthermore, it is concluded that the absence of callose deposition in BCA-infested leaves is not because of a stronger upregulation of callose-degrading beta-1,3-glucanases in this tissue, as compared to RWA-infested leaves.
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