Androgenesis-Based Doubled Haploidy: Past, Present, and Future Perspectives

Androgenesis, which entails cell fate redirection within the microgametophyte, is employed widely for genetic gain in plant breeding programs. Moreover, androgenesis-responsive species provide tractable systems for studying cell cycle regulation, meiotic recombination, and apozygotic embryogenesis within plant cells. Past research on androgenesis has focused on protocol development with emphasis on temperature pretreatments of donor plants or floral buds, and tissue culture optimization because androgenesis has different nutritional requirements than somatic embryogenesis. Protocol development for new species and genotypes within responsive species continues to the present day, but slowly. There is more focus presently on understanding how protocols work in order to extend them to additional genotypes and species. Transcriptomic and epigenetic analyses of induced microspores have revealed some of the cellular and molecular responses required for or associated with androgenesis. For example, microRNAs appear to regulate early microspore responses to external stimuli; trichostatin-A, a histone deacetylase inhibitor, acts as an epigenetic additive; ά-phytosulfokine, a five amino acid sulfated peptide, promotes androgenesis in some species. Additionally, present work on gene transfer and genome editing in microspores suggest that future endeavors will likely incorporate greater precision with the genetic composition of microspores used in doubled haploid breeding, thus likely to realize a greater impact on crop improvement. In this review, we evaluate basic breeding applications of androgenesis, explore the utility of genomics and gene editing technologies for protocol development, and provide considerations to overcome genotype specificity and morphogenic recalcitrance in non-model plant systems.

Bogus Data Acquisition Protocol I v1

The purpose of this protocol is to demo protocol development. The real version of this protocol might be developed with respect to existing publications. Here, I'll refer to publications using the Citation component.

Bogus Data Acquisition Protocol III v1

The purpose of this protocol is to demo protocol development.

Bogus data acquisition protocol II v1

The purpose of this protocol is to demo protocol development.

Bogus Sample Prep Protocol v1

This is a bogus protocol for assessing protocol development. Here, I'll use the citation component to refer to a publication that might influence protocol development.

Bogus Data Acquisition Protocol IV v1

The purpose of this protocol is to demo protocol development.

Modular automated bottom-up proteomic sample preparation for high-throughput applications v2

Manual proteomic sample preparation methods limit sample throughput and often lead to poor data quality when thousands of samples must be analyzed. Automated workflows are increasingly used to overcome these issues for some (or even all) of the sample preparation steps. Here, we detail three optimised step-by-step protocols to: (A) lyse Gram-negative bacteria and fungal cells; (B) quantify the amount of protein extracted; and (C) normalize the amount of protein and set up tryptic digestion. These protocols have been developed to facilitate rapid, low variance sample preparation of hundreds of samples, be easily implemented on widely-available Beckman-Coulter Biomek automated liquid handlers, and allow flexibility for future protocol development. By using this workflow 50 micrograms of peptides for 96 samples can be prepared for tryptic digestion in under an hour. We validate these protocols by analyzing 47 E. coli and R. toruloides samples and show that this modular workflow provides robust, reproducible proteomic samples for high-throughput applications. The expected results from these protocols are 94 peptide samples from Gram-negative bacterial and fungal cells prepared for bottom-up quantitative proteomic analysis without the need for desalting column cleanup and with peptide variance (CVs) below 15%.

Standardization of parenteral nutrition – a review of protocol development in a tertiary neonatal setup

Infants, who are either born preterm or with a weakened gastrointestinal tract, cannot tolerate feeds immediately after their birth normally through enteral route. This is where parenteral nutrition (PN) comes in, which employs ways to maintain the child’s nutrition through ways other than enteral route. PN can be of individualized type, one which is customized for each neonate, while the other is a standardized PN, which has some empirical components absolutely necessary for the infants. Both kinds help in maintaining the optimum nutritive requirements of the infants. However, there are some differences in their effects, which can be noteworthy.

PP296 Patient Involvement In An Assessment Of The Management Of Sudden Onset Severe Headache Presenting To The Emergency Department

IntroductionSudden onset severe headache is usually caused by a primary headache disorder but may be secondary to a more serious problem, such as subarachnoid hemorrhage (SAH). Very few patients who present to hospital with headache have suffered a SAH, but early identification is important to improve patient outcomes. A systematic review was undertaken to assess the clinical effectiveness of different care pathways for the management of headache, suspicious for SAH, in the Emergency Department. Capturing the perspective of patients was an important part of the research.MethodsThe project team included a patient collaborator with experience of presenting to the Emergency Department with sudden onset severe headache. Three additional patients were recruited to our advisory group. The patient's perspective was collected at various points through the project including at team meetings, during protocol development and when interpreting the results of the systematic review and drawing conclusions.ResultsPatients were reassured by the very high diagnostic accuracy of computed tomography (CT) for detecting SAH. Patients and clinicians emphasized the importance of shared decision making about whether to undergo additional tests to rule out SAH, after a negative CT result. When lumbar puncture was necessary, patients expressed a preference to have it on an ambulatory basis; further research on the safety and acceptability of ambulatory lumbar puncture was recommended.ConclusionsPatient input at the protocol development stage helped researchers understand the patient experience and highlighted important outcomes for assessment. Patient involvement added context to the review findings and highlighted the preferences of patients regarding the management of headache.

European NCAP Program Developments to Address Driver Distraction, Drowsiness and Sudden Sickness

Driver distraction and drowsiness remain significant contributors to death and serious injury on our roads and are long standing issues in road safety strategies around the world. With developments in automotive technology, including driver monitoring, there are now more options available for automotive manufactures to mitigate risks associated with driver state. Such developments in Occupant Status Monitoring (OSM) are being incorporated into the European New Car Assessment Programme (Euro NCAP) Safety Assist protocols. The requirements for OSM technologies are discussed along two dimensions: detection difficulty and behavioral complexity. More capable solutions will be able to provide higher levels of system availability, being the proportion of time a system could provide protection to the driver, and will be able to capture a greater proportion of complex real-word driver behavior. The testing approach could initially propose testing using both a dossier of evidence provided by the Original Equipment Manufacturer (OEM) alongside selected use of track testing. More capable systems will not rely only on warning strategies but will also include intervention strategies when a driver is not attentive. The roadmap for future OSM protocol development could consider a range of known and emerging safety risks including driving while intoxicated by alcohol or drugs, cognitive distraction, and the driver engagement requirements for supervision and take-over performance with assisted and automated driving features.