A rapid in vitro phenotypic assay of walnut shoots for pre-screening resistance to Phytophthora pini

2021 ◽  
Author(s):  
Paulo Adriano Zaini ◽  
Steven H. Lee ◽  
Charles Leslie ◽  
Sriema L. Walawage ◽  
Cai-Zhong Jiang ◽  
...  
Keyword(s):  
Phytophthora Species ◽  
In Vitro Assay ◽  
Crown Rot ◽  
Screening Methods ◽  
Breeding Programs ◽  
Vitro Assay ◽  
Horticultural Crops ◽  
Inoculation Procedure ◽  
Yield Quality

Phytophthora species cause crown, root, and aerial canker diseases in diverse horticultural crops, resulting in a loss of yield, quality and plant death. Breeding programs are interested in developing genotypes resistant to these and other pathogens, but current screening methods for tree crops are time-consuming, potentially spanning many years. Here we present a rapid in vitro assay in which walnut shoots obtained from tissue culture can be challenged with Phytophthora zoospores and monitored for symptom development within a week. As a proof of concept, two cultivars, scion variety ‘Chandler’ and clonal rootstock ‘RX1’, with known different degrees of crown rot susceptibility, were inoculated in vitro with P. pini (formerly included in P. citricola complex), and results of the assay reproducibly matched the differences in disease susceptibility generally observed for these two walnut selections under greenhouse and orchard conditions. In addition, when the in vitro assay was used for inoculations with P. capsici, which is not pathogenic on walnut, no disease development was observable, indicating that the assay can discriminate between species of Phytophthora that are aggressive on walnut and those not known as walnut pathogens. Our results suggest that this in vitro shoot inoculation procedure may provide rapid assessments useful for pre-screening resistance to Phytophthora in walnuts. Further testing of the assay is justified to determine whether it can resolve more subtle differences in resistance and whether it can be useful with other perennial hosts of Phytophthora.

In Vitro Assay of Platelet Adhesiveness with Washed and Tanned Human Red Cells

1968 ◽  
Vol 20 (03/04) ◽  
pp. 384-396 ◽  
Author(s):  
G Zbinden ◽  
S Tomlin
Keyword(s):  
Platelet Adhesion ◽  
Sodium Citrate ◽  
Blood Platelets ◽  
In Vitro Assay ◽  
Red Cells ◽  
Platelet Adhesiveness ◽  
Vitro Assay ◽  
In Vitro System ◽  
Human Red Cells

SummaryAn in vitro system is described in which adhesion of blood platelets to washed and tannic acid-treated red cells was assayed quantitatively by microscopic observation. ADP, epinephrine and TAME produced a reversible increase in platelet adhesiveness which was antagonized by AMP. With Evans blue, polyanetholsulfonate, phthalanilide NSC 38280, thrombin and heparin at concentrations above 1-4 u/ml the increase was irreversible. The ADP-induced increase in adhesiveness was inhibited by sodium citrate, EDTA, AMP, ATP and N-ethylmaleimide. EDTA, AMP and the SH-blocker N-ethylmaleimide also reduced spontaneous platelet adhesion to red cells. No significant effects were observed with adenosine, phenprocoumon, 5-HT, phthalanilide NSC 57155, various estrogens, progestogens and fatty acids, acetylsalicylic acid and similarly acting agents, hydroxylamine, glucose and KCN. The method may be useful for the screening of thrombogenic and antithrombotic properties of drugs.

Angiogenic Effect of Pravastatin Alone and with Sera from Healthy and Complicated Pregnancies Studied by in vitro Vasculogenesis/Angiogenesis Assay

2021 ◽  
pp. 1-9
Author(s):  
Anita Virtanen ◽  
Outi Huttala ◽  
Kati Tihtonen ◽  
Tarja Toimela ◽  
Tuula Heinonen ◽  
...  
Keyword(s):  
Direct Effect ◽  
Late Onset ◽  
Maternal Serum ◽  
In Vitro Assay ◽  
Serum Samples ◽  
Therapeutic Concentration ◽  
Tubule Formation ◽  
Vitro Assay ◽  
Angiogenesis Assay

<b><i>Objective:</i></b> To determine the direct effect of pravastatin on angiogenesis and to study the interaction between pravastatin and maternal sera from women with early- or late-onset pre-eclampsia (PE), intrauterine growth restriction, or healthy pregnancy. <b><i>Methods:</i></b> We collected 5 maternal serum samples from each group. The effect of pravastatin on angiogenesis was assessed with and without maternal sera by quantifying tubule formation in a human-based in vitro assay. Pravastatin was added at 20, 1,000, and 8,000 ng/mL concentrations. Concentrations of angiogenic and inflammatory biomarkers in serum and in test medium after supplementation of serum alone and with pravastatin (1,000 ng/mL) were measured. <b><i>Results:</i></b> Therapeutic concentration of pravastatin (20 ng/mL) did not have significant direct effect on angiogenesis, but the highest concentrations inhibited angiogenesis. Pravastatin did not change the levels of biomarkers in the test media. There were no changes in angiogenesis when therapeutic dose of pravastatin was added with maternal sera, but there was a trend to wide individual variation towards enhanced angiogenesis, particularly in the early-onset PE group. <b><i>Conclusions:</i></b> At therapeutic concentration, pravastatin alone or with maternal sera has no significant effect on angiogenesis, but at high concentrations the effect seems to be anti-angiogenic estimated by in vitro assay.

In Vitro Assay for Single-cell Characterization of Impaired Deformability in Red Blood Cells under Recurrent Episodes of Hypoxia

Lab on a Chip ◽  
2021 ◽  
Author(s):  
YUHAO QIANG ◽  
Jia Liu ◽  
Ming Dao ◽  
E Du
Keyword(s):  
Shear Stress ◽  
Red Blood Cells ◽  
Single Cell ◽  
Blood Cells ◽  
Blood Circulation ◽  
In Vitro Assay ◽  
Vitro Assay ◽  
Cyclic Shear

Red blood cells (RBCs) are subjected to recurrent changes in shear stress and oxygen tension during blood circulation. The cyclic shear stress has been identified as an important factor that...

Purification of Chalcone Synthase from Tulip Anthers and Comparison with the Synthase from Cosmos Petals

1981 ◽  
Vol 36 (1-2) ◽  
pp. 30-34 ◽  
Author(s):  
Rainer Sütfeld ◽  
Rolf Wiermann
Keyword(s):  
Chalcone Synthase ◽  
In Vitro Assay ◽  
Flavonoid Biosynthesis ◽  
Chalcone Isomerase ◽  
Gel Chromatography ◽  
Purification Procedure ◽  
Reaction Products ◽  
Complete Elimination ◽  
Vitro Assay

Abstract Chalcone synthase was isolated from both anthers of Tulipa cv. “Apeldoorn” and petals of Cosmos sulphureus Cav. After certain prepurification steps, the enzymes were further purified using gel chromatography on Sephadex G-200 followed by repeated hydroxylapatite absorption chromatography. Both the enzymes showed the same chromatographic properties. After gel chromatography as well as after the first hydroxylapatite fractionation, the reaction products appeared as flavanones. However, after the second hydroxylapatite step, production of chalcones was observed. Like the enzyme from tulip anthers, the synthase from Cosmos petals produced the correspondingly substituted chalcones when p-coumaroyl-CoA, caffeoyl-CoA and feruloyl-CoA, respectively, were used as substractes. In both the cases, the ratios of the different chalcones produced were found to be about the same. The appearance of chalcone synthesis in this in vitro assay is caused by the complete elimination of chalcone isomerase in the purification procedure. The importance of the isomerase for flavonoid biosynthesis, particularly in plant systems which are accumulating chalcones, is discussed.

scholarly journals In Vitro Characterization of the Enzyme Properties of the Phospholipid N-Methyltransferase PmtA from Agrobacterium tumefaciens

2009 ◽  
Vol 191 (7) ◽  
pp. 2033-2041 ◽  
Author(s):  
Meriyem Aktas ◽  
Franz Narberhaus
Keyword(s):  
Agrobacterium Tumefaciens ◽  
In Vitro Assay ◽  
Enzyme Properties ◽  
Vitro Assay ◽  
Plant Infection ◽  
In Vitro Characterization ◽  
End Products ◽  
And Control

ABSTRACT Agrobacterium tumefaciens requires phosphatidylcholine (PC) in its membranes for plant infection. The phospholipid N-methyltransferase PmtA catalyzes all three transmethylation reactions of phosphatidylethanolamine (PE) to PC via the intermediates monomethylphosphatidylethanolamine (MMPE) and dimethylphosphatidylethanolamine (DMPE). The enzyme uses S-adenosylmethionine (SAM) as the methyl donor, converting it to S-adenosylhomocysteine (SAH). Little is known about the activity of bacterial Pmt enzymes, since PC biosynthesis in prokaryotes is rare. In this article, we present the purification and in vitro characterization of A. tumefaciens PmtA, which is a monomeric protein. It binds to PE, the intermediates MMPE and DMPE, the end product PC, and phosphatidylglycerol (PG) and phosphatidylinositol. Binding of the phospholipid substrates precedes binding of SAM. We used a coupled in vitro assay system to demonstrate the enzymatic activity of PmtA and to show that PmtA is inhibited by the end products PC and SAH and the antibiotic sinefungin. The presence of PG stimulates PmtA activity. Our study provides insights into the catalysis and control of a bacterial phospholipid N-methyltransferase.

scholarly journals DIGESTIBLE ENERGY IN FORAGE BYIN VIVOANDIN VITROASSAYS

1970 ◽  
Vol 50 (3) ◽  
pp. 557-562 ◽  
Author(s):  
J. E. TROELSEN
Keyword(s):  
Energy Analysis ◽  
Energy Content ◽  
In Vitro Assay ◽  
Pure Species ◽  
Vitro Assay ◽  
Digestible Energy ◽  
The Relationship ◽  
Cattle And Sheep

Forage of six pure species was harvested for hay at several maturity stages during four years. The digestible energy content of 102 different lots of hay was determined by feeding to four groups of sheep during the same period, and by in vitro digestions and energy analysis of the undigested residues. The relationship between digestible energy content assayed by the two methods was highly significant (r = 0.85) and did not differ between years and species. Exclusion from regression of the hays containing less than 2 or more than 3 digestible kcal/g revealed that the in vitro assay could reproduce the in vivo digestible energy value with a standard deviation of 0.31 in over 70% of the hays. This represented the maturity and quality range of forage commonly fed to cattle and sheep. The in vitro assay therefore appeared promising for commercial quality determinations.

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