Commun Biol.2020 Dec 10;3(1):750.doi: 10.1038/s42003-020-01480-5.
Biological reagents and resources. The RHΔku80Δhxgprt 75 and
RHΔku80Δhxgprt-TIR1 49 strains of T. gondii were kindly offered by Vern Carruthers (University of Michigan, MI) and David Sibley (Washington State University, St. Louis, MO), respectively. The Δtgcds1 r mutant was generated in our earlier study 7 . The pG140 plasmid was donated by Markus Meissner (Ludwig-Maximilians University, Munich). Antibodies recognizing the TgHSP90, TgGAP45,TgPDH-E1a, TgSAG2, and TgIMC3 proteins were bestowed by Sergio Angel (IIB-INTECH, Argentina), Dominique Soldati-Favre (University of Geneva, Switzer-land), Bang Shen (Huazhong Agricultural University, China), Honglin Jia (HarbinVeterinary Research Institute, China) and Marc-Jan Gubbels (Boston College,MA), respectively. Anti-TgSERCA and anti-TgPSD1mt antisera were obtained from FriendBio Bioscience and Technology (China). Other primary antibodies recognizing the TgSAG1 protein and engineered epitopes (HA and Ty1) were obtainedfrom ThermoFisher Scientific and Sigma-Aldrich (Germany). Corresponding secondary antibodies (Alexa488, Alexa594) were procured from Life Technologies(Germany). [ 3 H]-myo-inositol, and silica plates for TLC were obtained from American Radiolabeled Chemicals (St. Louis, MO) and Merck Millipore (Billerica,Fig. 9 Model of PtdIns biosynthesis and homeostasis of anionic phospholipids in T. gondii. PtdIns is made by TgPIS in the Golgi network using CDP-DAGand myo-inositol. CDP-DAG is generated by TgCDS1 in the endoplasmic reticulum, whereas myo-inositol is imported from the milieu. De novo synthesis ofmyo-inositol involves conversion of glucose to glucose-6-phosphate by hexokinase, followed by the action of inositol-3P synthase to produce inositol-3-phosphate. Successively, Ins3P is dephosphorylated to make inositol by inositol-3P monophosphatase. There is no biochemical and genetic evidence for the latter enzyme in T. gondii. Knockdown of TgPIS reduces the level of many PtdIns species along with homeostatic modulation of other anionic lipids (namelyPtdThr, PtdSer, PtdGro species). PtdIns species with shorter chains (e.g., C30/C32/C34) are synthesized de novo, while PtdIns species with relatively longer acyl chain (C38/C40) are salvaged by the parasite from its host cell. Based on isotope labeling studies, we speculate that both routes contribute to the synthesis of C36 PtdIns species (not shown). Transporters, inter-organelle contact sites, or other lipid trafficking pathways, as stated, enable transport of myo-inositol and CDP-DAG. The question-marked reaction specifies yet-unknown enzyme involved in CDP-DAG-dependent PtdSer synthesis. Selected abbreviations: CDS, CDP-DAG synthase; PGPS, PtdGro phosphate synthase; PGPP, PtdGro phosphate phosphatase; PSS, PtdSer synthase; PTS, PtdThrsynthase; PPM, parasite’s plasma membrane; PVM, parasitophorous vacuole membrane.
MA), respectively. [ 13 C]-myo-inositol was kindly provided by Dorothea Fiedler(Leibniz Research Institute, Berlin). Lipid standards and 1,2-dioleoyl-sn-glycero-3-(cytidine diphosphate) (CDP-DAG) were delivered by Avanti Polar Lipids (USA).DNA oligonucleotides (Supplementary Table 2) were purchased from Life Technologies.
S Rong, Y Zhou, M Niu, Q Li, M Wang… - … Material Science …, 2016 - atlantis-press.com
… Anti-VP1 (GI.1) or anti-VP1 (GII.4) polyclonal antisera (FriendbioScience and Technology (Wuhan) Co., Ltd., Hubei, China) were used as primary antibodies. The horseradish peroxidase-conjugated goat anti-mouse immunoglobulin G (IgG) was used as secondary antibodies …
J Wu, W Deng, D Lin, X Deng, Z Ma - Journal of agricultural and …, 2018 - ACS Publications
Production of Rabbit Polyclonal Antibodies. In order to generate antibodies against Cit s 1.01, Cit s 2.01, and Cit s 3.01, two New Zealand white rabbits (purchased from Hubei Provincial Center for Disease Control and Prevention, China) were immunized subcutaneously with 0.1 mg of recombinant GST-(Cit s 1.01)×2, GST-(Cit s 2.01)×3, and Trx-His-(Cit s 3.01)×2 proteins, respectively. Immunization and serum collection were performed by Friendbio Science & Technology (Wuhan) Co., Ltd. The protocol was approved by the Institute of Animal Care and Use Committee (no.201703083), according to national guidelines for animal care and handling, China. Immunization was performed every 2 weeks for a period of 3 months. The sera were collected after 3 months of immunization. Antibody titers were measured by ELISA. The sera were stored at −80 °C for further use
RH Gan, Y Wang, Z Li, ZX Yu, XY Li… - Molecular Biology …, 2021 - academic.oup.com
Polyclonal antibody preparation, Western blot, histological section and immunofluorescence Polyclonal antibodies respectively specific to the CgFoxl2a and CgFoxl2b were produced by Friendbio Science & Technology (Wuhan) Co,.Ltd. Western blot,histological section and immunofluorescence were performed as described previously (Liu et al. 2015; Yang et al. 2017), and the results were acquired with ImageQuant LAS 4000mini (GE) and Carl Zeiss upright fluorescence microscope Axio Imager M2 (Analytical & Testing Center, IHB, CAS), respectively.