Comprehensive solution for algal phenotype research

Algae are a general term for a series of aquatic organisms such as cyanobacteria, diatoms, dinoflagellates, green algae, brown algae, and red algae. There are various forms of microalgae, ranging from micrometer sized single celled microalgae to large brown algae that can grow up to several meters or even tens of meters in length. Algae, as the most important primary producers in water bodies, play an extremely important role in the stability of the entire ecosystem and even the Earth's sphere. Model algae such as Chlamydomonas reinhardtii and cyanobacteria provide excellent materials for research on functional genes, biological evolution, photosynthesis, and more. Meanwhile, many economic algae also play important roles in industries such as food, medicine, and energy. Harmful ecological phenomena such as algal blooms and red tides are also caused by algae. Therefore, the study of algae has always been a very important hotspot in biology and ecology

At present, one of the most eye-catching research hotspots in the field of life sciences - phenotype omics research, is mainly focused on the field of plants/crops. In the field of algae, phenotype omics research has just begun, but the development speed is extremely rapid. On the one hand, algal genomics is a supplement and validation of algal genomics, explaining the complex roles and intermediate processes of genome and environmental factors in plant phenotypes; On the other hand, it can be used for stress resistance physiology research and genetic breeding of economic algae, comprehensively analyzing algal phenotypes and obtaining better economic algae species; It can also be used for studying the occurrence mechanism of algal blooms and red tides.

Algae phenotype omics requires a comprehensive analysis of the phenotypic characteristics of algae, especially the measurement and analysis of photosynthetic physiology, morphology, color, pigment composition and distribution, photosynthetic contributions of different pigments, stress physiology, etc., in order to digitize algal phenotypes, visualize physiological ecology and functions. This requires technical solutions specifically designed for algal phenotypes.

The instruments listed in the plan can be flexibly combined according to specific research needs, and all instruments have high international recognition. Scientists at home and abroad have published a large number of scientific papers using these instruments. Below are some application cases of system solutions.

1. In depth study on the mechanism of photosynthesis in blue-green algae

Researcher Wang Qiang from the Institute of Hydrobiology, Chinese Academy of Sciences, studied the effect of nitrite stress on blue-green algaeSynechocystisThe stress mechanism of sp. PCC 6803 photosystem II, usingAquaPenHandheld algae fluorescence measuring instrumentFL3500Chlorophyll fluorescence analyzer (models before FL6000) and TL chlorophyll thermoluminescence system were used to measure and analyze the maximum photochemical efficiency of Fv/Fm, OJIP rapid fluorescence kinetics curve, QA re oxidation kinetics curve, S-state measurement, and TL thermoluminescence curve, respectively. The study ultimately proves that nitrite stress first affectsSynechocystisSp. PCC 6803 photosystem II receptor side (Zhan X, 2017)

Left: Experimental sample; In the middle:Fv/FmRight: OJIP rapid fluorescence kinetics curve

Left: QA reoxygenation kinetics curve; Middle: S-state measurement: Right: TL thermoluminescence curve

Left: AquaPen handheld algae fluorescence measuring instrument; Chinese: FL3500 dual modulation chlorophyll fluorescence analyzer; Right: TL Chlorophyll Thermoluminescence System

reference: Zhang X, Ma F, Zhu X, Zhu J, Rong J, Zhan J, Chen H, He C, Wang Q. 2017. The acceptor side of photosystem II is the initial target of nitrite stress in Synechocystis sp. strain PCC 6803. Appl Environ Microbiol 83:e02952-16

2. Multi omics analysis of molecular events in Chlamydomonas reinhardtii

Chlamydomonas reinhardtiiChlamydomonas reinhardtiiMetabolism has good adaptability to environmental changes. Daniela Strenkert from the University of California attempted to simulate a light dark cycle and study the transcriptome and proteomic changes of Chlamydomonas reinhardtii during that day. This study first requires the ability to accurately simulate a dynamically changing cultivation environment, while also monitoring the relevant cultivation environment, algal density, physiology, and other factors. This can provide the most accurate samples for subsequent omics analysis. At present, the only algae cultivation monitoring instrument that can meet this requirement is the FMT150 algae cultivation and online monitoring system. The cultivation conditions are set as follows:

nSimultaneous cultivation of multiple FMT150 machines

n200 μERed/blue light, 12 hour day night alternation, simulating sunrise and sunset

n28 ℃ during the day and 18 ℃ at night

nVentilation rate 0.2 L/min

nReal time monitoring of temperature, OD680, and pH

Pre culture with constant turbidity for 5 days, control OD680 to 0.4

Left: A.FMT150 schematic diagram, B-D. Real time monitoring of temperature, OD680, and pH during the cultivation process; Right:Dynamic variation of maximum photochemical efficiency Fv/Fm under different light intensities

Chlorophyll fluorescence analysis is an indispensable part of photosynthetic physiology and phenotype omics research. Therefore, this study simultaneously usedFluorCamMeasurement of the dynamic changes in the maximum photochemical efficiency Fv/Fm of Chlamydomonas reinhardtii under different light intensities using a closed fluorescence imaging system, reflecting the effect of light intensity on Chlamydomonas reinhardtii and conducting correlation analysis with proteomic and transcriptome data

Left:FMT150Algae cultivation and online monitoring system; Right: FluorCam Closed Chlorophyll Fluorescence Imaging System

reference: Strenkert D,et al.2019, Multiomics resolution of molecular events during a day in the life ofChlamydomonas. PNAS 116 (6): 2374-2383

3. Monitoring the Climate Response of Antarctic Algae and Lichens

2006In, the Czech Republic built the Johann Gregor Mendel station on James Ross Island in Antarctica. Researchers from Masaryk University in the Czech Republic stationed at the station have been studying the response of local algae and lichens to rising temperatures in Antarctica since 2007, in order to assess the impact of greenhouse effects on the Antarctic ecosystem. At that time, they used the AquaPen/FluorPen series handheld chlorophyll fluorescence measuring instrument specifically designed to enhance polar adaptability to detect the photosynthetic physiology and growth status of algae and lichens. The AquaPen/FluorPen can be manually operated and also has the function of unmanned monitoring of chlorophyll fluorescence, performing well in the harsh environment of Antarctica.

In recent years, researchers have started using Monitoring Pen chlorophyll fluorescence automatic monitoring devices specifically designed for monitoring experiments. The Monitoring Pen can automatically work continuously for 2 years under ideal conditions, equipped with two versions: land enhanced version and underwater enhanced version.

Left: AquaPen/FluorPen used by Johann Gregor Mendel station from 2007 to 2009; Right: Monitoring Pen used in recent years

Left: Monitoring Pen Land Enhanced Edition; Right: Monitoring Pen Underwater Enhanced Edition

Reference: Bart á k M,et al.2009, Long-term study on vegetation responses to manipulated warming using open top chambers installed in three contrasting Antarctic habitats. Structure and function of antarctic terrestrial ecosystems, Masaryk University

4. Molecular mechanism of response to high light stress in Chlamydomonas reinhardtii

All photosynthetic organisms must cope with excessive light exposure to avoid photosynthetic oxidative stress. For plants and green algae, the fastest response mechanism to high light is non photochemical quenching (NPQ) of photosystem II. This process allows photosystem II to safely dissipate excess energy in the form of heat. PsbS protein is an important sensor in this process.

To determine the presence of PsbS protein in Chlamydomonas reinhardtiiChlamydomonas reinhardtiiThe role of NPQ and photoprotection in the cultivation of algae or Arabidopsis thaliana by Tibiletti T et al. from the University of Aix MarseillepsbSThe chloroplast transgenic strain of the gene. The NPQ imaging analysis using the FluorCam open chlorophyll fluorescence imaging system ultimately showed that both PsbS proteins can enhance the wild-type and wild-type of Chlamydomonas reinhardtiinpq4The NPQ of the mutant strain, but no clear photoprotective activity was observed through Fv/Fm measurement.

Left: NPQ imaging image; Medium: Dynamic changes in NPQ during light dark cycles; Right: Fv/Fm data analysis

At the same time, they also used two white LED light source boards to simulate sustained high light stress (1200 µ mol m-2 s-1); Fytoled light source system was used to simulate fluctuating lighting conditions (3 minutes 1200 µ mol m-2 s-1+3 minutes 45 µ mol m-2 s-1) for growth rate experiments; The FluorCam system comes with a blue LED light source board to simulate fluctuating lighting conditions (3 minutes 1200 µ mol m-2 s-1+3 minutes 20 µ mol m-2 s-1) for Fv/Fm analysis. These LED light source boards actually come from the same technology source, namely PSI's SL3500 LED light source board. The development of this LED light source is aimed at providing high-intensity, high uniformity, and high-purity illumination for the FluorCam system to measure chlorophyll fluorescence imaging, while also equipped with microsecond level automatic control function. The cultivation system developed based on this LED light source naturally possesses high performance that other cultivation systems cannot achieve. Many researchers in Europe and America use this light source to build their own designed cultivation systems. For example, in the article "Recovery rates reflect distance to a tipping point in a living system" published in Nature in 2012, the author used the SL3500 light source to build a self-designed algae cultivation system.

Left: FluorCam Open Chlorophyll Fluorescence Imaging System; Chinese: SL3500 LED light source board; Right: AlgaeTron/FytoScope Algae/Plant Incubator

reference: Tibiletti T,et al. 2016. Chlamydomonas reinhardtiiPsbS protein is functional and accumulates rapidly and transiently under high light. Plant Physiology, 171(4): 2717-2730