The Dawn of Plant Molecular Biology: How Three Key Methodologies Paved the Way

Somssich, Marc

doi:10.1002/cpz1.417

Cited by 6 publications

(8 citation statements)

References 221 publications

(207 reference statements)

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“…Since the development of molecular cloning in the early 1970s, the isolation of genes and promoters, and subsequent transgenesis of model organisms, has become standard practice in the life sciences (Somssich, 2022). The development of more advanced cloning methods, particularly the recombination-based Gateway technology in the year 2000, made the creation of expression clones ready for transformation ever easier (Hartley et al ., 2000).…”

Section: Introductionmentioning

confidence: 99%

pGG-PIP: A GreenGate (GG) entry vector collection with Plant Immune system Promoters (PIP)

Calabria

Rast-Somssich

Wang

et al. 2022

Preprint

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The regulatory sequences controlling the expression of a gene (i.e. the promoter) are essential to properly understand a gene's function. From their use in mutant complementation assays, to studying their responsiveness to different stimuli via transcriptional reporter lines or using them as proxy for the activation of certain pathways, assays using promoter sequences are valuable tools for insight into the genetic architecture underlying plant life. The GreenGate (GG) system is a plant-specific variant of the Golden Gate assembly method, a modular cloning system that allows the hierarchical assembly of individual donor DNA fragments into one expression clone via a single reaction step. The assembly is based on specific recognition sites for the individual donor fragments, derived i.e., from a promoter, coding sequence, resistance gene or a protein tag. Here, we present a collection of 75 GG entry vectors carrying putative regulatory sequences for Arabidopsis thaliana genes involved in many different pathways of the plant immune system, designated Plant Immune system Promoters (PIP). This pGG-PIP entry vector set enables the rapid assembly of expression vectors to be used for transcriptional reporters of plant immune system components, mutant complementation assays when coupled with coding sequences, mis-expression experiments for genes of interest, or the targeted use of CRISPR/Cas9 genome editing. We used pGG-PIP vectors to create fluorescent transcriptional reporters in A. thaliana and demonstrated the potential of these reporters to image the responsiveness of specific plant immunity genes to infection and colonization by the fungal pathogen Fusarium oxysporum. Using the PLANT ELICITOR PEPTIDE (PEP) pathway as an example, we show this pathway is locally activated in response to colonization by the fungus.

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Section: Introductionmentioning

confidence: 99%

pGG-PIP: A GreenGate (GG) entry vector collection with Plant Immune system Promoters (PIP)

Calabria

Rast-Somssich

Wang

et al. 2022

Preprint

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“…One of the first labs in the plant field to adopt the CLSM was the group of Elliott Meyerowitz, who had already been instrumental in pioneering A. thaliana as a plant model (Somssich, 2018(Somssich, , 2022. In the early 1990s, Mark Running from the Meyerowitz lab developed CLSM to image Arabidopsis meristems, using propidium iodide as a marker for nuclei (Clark, Running, & Meyerowitz, 1993;Running, Clark, & Meyerowitz, 1995).…”

Section: Confocal Microscopy (1967-1985)mentioning

confidence: 99%

“…The A. thaliana data was only featured in his complete thesis, however, and was omitted from the journal publication, as it was not regarded as important enough at the time (Somssich, 2018). His observation that A. thaliana carries only five chromosomes was among the reasons Laibach later proposed this little weed as a plant model organism, thereby helping to change this view (Somssich, 2018(Somssich, , 2022. Following this work, Emil Heitz analyzed the chromosomes of liverworts in closer detail, thereby following the footsteps of both M. polymorpha pioneer Brisseau de Mirbel and A. thaliana pioneer Laibach (Berger, 2019;Heitz, 1928).…”

Section: The Cell Theory Cell Division and Plant Cell Chromosomes (18...mentioning

confidence: 99%

“…Later on, more dyes for specific structures and organelles were added to the toolkit, such as 3,3 -dihexyloxacarbocyanine iodide (DiOC6(3)) to mark the plant endoplasmic reticulum (Quader & Schnepf, 1986). Furthermore, with the adoption of A. thaliana as plant model organism and the establishment of plant transformation, the field of molecular biology had finally reached the plant sciences, and with it brought the first genetically encoded reporter for plant light microscopy (Jefferson, Kavanagh, & Bevan, 1987;Somssich, 2018Somssich, , 2019Somssich, , 2022. This came in the form of the Escherichia coli β-glucuronidase (GUS) gene (Jefferson et al, 1987).…”

Section: Differential Interference Contrast Microscopy (1955)mentioning

confidence: 99%

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A Short History of Plant Light Microscopy

Somssich

2022

Current Protocols

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When the microscope was first introduced to scientists in the 17 th century it started a revolution. Suddenly a whole new world, invisible to the naked eye, opened up to curious explorers. In response to this realization Nehemiah Grew, one of the early microscopists, noted in 1682 'that Nothing hereof remains further to be known, is a Thought not well Calculated.' 1 . And indeed, with ever increasing resolution, there really does not seem to be an end to what can be explored with a microscope. The Beginnings: Plant Internal Structures and 'Cells' (1600-1835)While simple lenses were being used as magnifying glasses for several centuries, the early 17 th century brought the invention of the compound microscope, and with it launched the

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“…Arabidopsis thaliana ( Arabidopsis ) has been used as a model plant species for developmental biology, physiology, and molecular genetics studies since the 1980s due to its unparalleled intrinsic and experimental properties (Meyerowitz & Pruitt, 1985; Somssich, 2022). It has also become a model for cytogenetic studies with only five pairs of chromosomes, and the first light microscopic atlas of the meiosis of Arabidopsis was published in 1996 (Ross, Fransz, & Jones, 1996).…”

Section: Introductionmentioning

confidence: 99%

Evaluation of Crossover Number, Distribution, and Interference Using Cytological Assays in Arabidopsis

Huang

Wang

2022

Current Protocols

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Meiosis involves the replication of nuclear chromosomes in a parent cell, followed by two successive nuclear divisions to produce haploid spores, which differentiate into the gametophyte generations that produce the egg and sperm in plants. Meiotic recombination or crossover (CO) is a hallmark of meiosis that allows shuffling of genetic information between homologous chromosomes (homologs), thereby giving rise to genetically diverse progeny cells and, ultimately, individuals in the progeny; this opens vast opportunities for genetic differentiation and hence speciation. Meiotic COs also ensure the formation of bivalents and fidelity of their equal segregation. Therefore, mechanisms that regulate meiotic recombination have been extensively studied in multiple species. Several approaches have been developed to observe or estimate the frequency of CO, in which CO can be visualized and analyzed cytologically by estimating the number of chiasma (plural chiasmata), a position where nonsister chromatids exchange genetic material between homologs. Furthermore, one CO event might influence the occurrence of another one nearby, along a chromosome; this is known as CO interference. Over the past decades, visualizing CO events and measuring CO interference have contributed greatly to our understanding of the regulatory mechanisms of meiotic recombination. Here, we describe protocols to estimate the number of chiasmata and CO interference in Arabidopsis using cytological methods involving chromosome spreads and immunostaining. Specifically, we describe how chromosome spreads can be used to estimate the number of chiasmata based on the conformations of metaphase I bivalents and provide a revised acid-based quick immunostaining assay that permits high-throughput and quantitative digital estimation of the relative distance between adjacent interference-sensitive CO foci at diakinesis. These methods can be easily established or modified, if necessary, for studying meiotic recombination in other plants and crops.

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The Dawn of Plant Molecular Biology: How Three Key Methodologies Paved the Way

Cited by 6 publications

References 221 publications

pGG-PIP: A GreenGate (GG) entry vector collection with Plant Immune system Promoters (PIP)

pGG-PIP: A GreenGate (GG) entry vector collection with Plant Immune system Promoters (PIP)

A Short History of Plant Light Microscopy

Evaluation of Crossover Number, Distribution, and Interference Using Cytological Assays in Arabidopsis

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