Genome-Wide High-Throughput RNAi Screening for Identification of Genes Involved in Protein Production.

With an increasing number of blockbuster drugs being recombinant mammalian proteins, protein production platforms that focus on mammalian proteins have had a profound impact in many areas of basic and applied research. Many groups, both academic and industrial, have been focusing on developing cost-effective methods to improve the production of mammalian proteins that would support potential therapeutic applications. As it stands, while a wide range of platforms have been successfully developed for laboratory use, the majority of biologicals are still produced in mammalian cell lines due to the requirement for posttranslational modification and the biosynthetic complexity of target proteins.

Immune system modulation & virus transmission during parasitism identified by multi-species transcriptomics of a declining insect biocontrol system.

Background: The Argentine stem weevil (ASW, Listronotus bonariensis) is a significant pasture pest in Aotearoa New Zealand, primarily controlled by the parasitoid biocontrol agent Microctonus hyperodae. Despite providing effective control of ASW soon after release, M. hyperodae parasitism rates have since declined significantly, with ASW hypothesised to have evolved resistance to its biocontrol agent.

Chromosome-level genome assemblies of two parasitoid biocontrol wasps reveal the parthenogenesis mechanism and an associated novel virus.

Background: Biocontrol is a key technology for the control of pest species. Microctonus parasitoid wasps (Hymenoptera: Braconidae) have been released in Aotearoa New Zealand as biocontrol agents, targeting three different pest weevil species. Despite their value as biocontrol agents, no genome assemblies are currently available for these Microctonus wasps, limiting investigations into key biological differences between the different species and strains.

The venom composition and parthenogenesis mechanism of the parasitoid wasp Microctonus hyperodae, a declining biocontrol agent.

A biocontrol system in New Zealand using the endoparasitoid Microctonus hyperodae is failing, despite once being one of the most successful examples of classical biocontrol worldwide. Though it is of significant economic importance as a control agent, little is known about the genetics of M. hyperodae.

Evaluation protocol for CRISPR/Cas9-mediated CD19 knockout GM24385 cells by flow cytometry and Sanger sequencing.

Although several genome editing options are available, CRISPR/Cas9 is one of the most commonly used systems for protein and advanced therapies. There are some long-term data regarding genomic and phenotypic stability, however, information is sparse. Flow cytometry can offer a method to characterize these edited cells for longitudinal studies.

Towards Quantitative and Standardized Serological and Neutralization Assays for COVID-19.

Quantitative and robust serology assays are critical measurements underpinning global COVID-19 response to diagnostic, surveillance, and vaccine development. Here, we report a proof-of-concept approach for the development of quantitative, multiplexed flow cytometry-based serological and neutralization assays. The serology assays test the IgG and IgM against both the full-length spike antigens and the receptor binding domain (RBD) of the spike antigen.

Stochastic Reaction-Diffusion Model of the Binding of Monoclonal Antibodies to CD4 Receptors on the Surface of T Cells.

A stochastic reaction-diffusion model was developed to describe the binding of labeled monoclonal antibodies (mAbs) to CD4 receptors on the surface of T cells. The mAbs diffused to, adsorbed on, and underwent monovalent and bivalent binding to CD4 receptors on the cell surface. The model predicted the time-dependent nature of all populations involved in the labeling process.

Genetic Diversity in Invasive Populations of Argentine Stem Weevil Associated with Adaptation to Biocontrol.

Modified, agricultural landscapes are susceptible to damage by insect pests. Biological control of pests is typically successful once a control agent has established, but this depends on the agent's capacity to co-evolve with the host. Theoretical studies have shown that different levels of genetic variation between the host and the control agent will lead to rapid evolution of resistance in the host.

TrichoGate: An Improved Vector System for a Large Scale of Functional Analysis of Genes.

Species of the genus are ubiquitous in the environment and are widely used in agriculture, as biopesticides, and in the industry for the production of plant cell wall-degrading enzymes. represents an important genus of endophytes, and several species have become excellent models for the study of fungal biology and plant-microbe interactions; moreover, are exceptional biotechnological factories for the production of bioactive molecules useful in agriculture and medicine. Next-generation sequencing technology coupled with systematic construction of recombinant DNA molecules provides powerful tools that contribute to the functional analysis of genetics, thus allowing for a better understanding of the underlying factors determining its biology.

Improved protein expression in HEK293 cells by over-expressing miR-22 and knocking-out its target gene, HIPK1.

Stable cell lines can continuously produce a recombinant protein without the need to repeatedly engineer the genome. In a previous study HIPK1, Homeodomain-interacting Protein Kinase 1, was found to be a target of the microRNA miR-22 that, when repressed, improved expression of both an intracellular and a secreted protein. In this report, HEK293 cells stably over-expressing miR-22 were compared with HEK293 with knockout of HIPK1, executed by CRISPR/Cas9, for their ability to improve recombinant protein expression.

Fed-batch high-cell-density fermentation strategies for Pichia pastoris growth and production.

Pichia pastoris is extensively used to produce various heterologous proteins. Amounts of biopharmaceutical drugs and industrial enzymes have been successfully produced by fed-batch high-cell-density fermentation (HCDF) of this cell factory. High levels of cell mass in defined media can be easily achieved and therefore large quantities of recombinant proteins with enhanced activities and lower costs can be obtained through HCDF technology.

Genome-Wide High-Throughput RNAi Screening for Identification of Genes Involved in Protein Production.

With an increasing number of blockbuster drugs being recombinant mammalian proteins, protein production platforms that focus on mammalian proteins have had a profound impact in many areas of basic and applied research. Many groups, both academic and industrial, have been focusing on developing cost-effective methods to improve the production of mammalian proteins that would support potential therapeutic applications. As it stands, while a wide range of platforms have been successfully developed for laboratory use, the majority of biologicals are still produced in mammalian cell lines due to the requirement for posttranslational modification and the biosynthetic complexity of target proteins.

Continuous production process of retroviral vector for adoptive T- cell therapy.

Adoptive T-Cell therapy is being considered as a promising method for cancer treatment. In this approach, patient's T cells are isolated, modified, expanded, and administered back to the patient. Modifications may include adding specific T cell receptors (TCR) or chimeric antigen receptors (CAR) to the isolated cells by using retroviral vectors.

Methods for Using Small Non-Coding RNAs to Improve Recombinant Protein Expression in Mammalian Cells.

The ability to produce recombinant proteins by utilizing different "cell factories" revolutionized the biotherapeutic and pharmaceutical industry. Chinese hamster ovary (CHO) cells are the dominant industrial producer, especially for antibodies. Human embryonic kidney cells (HEK), while not being as widely used as CHO cells, are used where CHO cells are unable to meet the needs for expression, such as growth factors.

Identifying HIPK1 as Target of miR-22-3p Enhancing Recombinant Protein Production From HEK 293 Cell by Using Microarray and HTP siRNA Screen.

Protein expression from human embryonic kidney cells (HEK 293) is an important tool for structural and clinical studies. It is previously shown that microRNAs (small, noncoding RNAs) are effective means for improved protein expression from these cells, and by conducting a high-throughput screening of the human microRNA library, several microRNAs are identified as potential candidates for improving expression. From these, miR-22-3p is chosen for further study since it increased the expression of luciferase, two membrane proteins and a secreted fusion protein with minimal effect on the cells' growth and viability.

Factors controlling sediment denitrification in midwestern streams of varying land use.

We investigated controls on stream sediment denitrification in nine headwater streams in the Kalamazoo River Watershed, Michigan, USA. Factors influencing denitrification were determined by using experimental assays based on the chloramphenicol-amended acetylene inhibition technique. Using a coring technique, we found that sediment denitrification was highest in the top 5 cm of the benthos and was positively related to sediment organic content.

Functional identification of an Arabidopsis pectin biosynthetic homogalacturonan galacturonosyltransferase.

Galacturonosyltransferases (GalATs) are required for the synthesis of pectin, a family of complex polysaccharides present in the cell walls of all land plants. We report the identification of a pectin GalAT (GAUT1) using peptide sequences obtained from Arabidopsis thaliana proteins partially purified for homogalacturonan (HG) alpha-1,4-GalAT activity. Transient expression of GAUT1 cDNA in the human embryonic kidney cell line HEK293 yielded uridine diphosphogalacturonic acid:GalAT activity.