Plant Symposia and Workshops - Society for In Vitro Biology
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In Vitro Cellular & Developmental Biology - Animal
https://doi.org/10.1007/s11626-020-00464-3
2020 WORLD CONGRESS ON IN VITRO BIOLOGY ABSTRACT ISSUE
Plant Symposia and Workshops
P-1 P-2
Conservation of Magnolia Spp Using Cryobiotechnology: Bumps, Potholes and Crashes Along the Way to Successful
From Wild Collection to Ex Vitro Hardening. R. FOLGADO. Micropropagation: Prevention and Intervention. C. J. SLUIS,
The Huntington Library, Art Museum, and Botanical Gardens, C. Del Cid, G. Y. Orozco, and B. Gytri. Tissue-Grown
1151 Oxford Rd., San Marino, CA 91108. Email: Corporation, 15245 W. Telegraph Road, Santa Paula,
rfolgado@huntington.org CA 93060. Email: carolynsluis@gmail.com,
celiadelcid@tissuegrown.com
The magnolias are trees or shrubs which are broadly appreciated
as ornamental plants around the world; they are also used as me- The scale-up of tissue cultured plants from a few hundred test
dicinal plants and for timber in their local communities. There are tubes to a productive base of thousands of micropropagated
concerns about the conservation of half of the Magnolia species, units ready for the greenhouse, presents a unique set of chal-
according to the Red List of Magnoliaceae (Rivers et al., 2016). lenges to the commercial micropropagation specialist.
Criobiotechnology tools, including tissue culture and cryopreser- Processes and protocols which work well at the level of a few
vation (i.e. storage at ultra-low temperatures), can help to ensure dozen test tubes, can perform differently when cultured in
the ex situ preservation of trees that produce seeds with low- higher numbers and in larger vessels. Even when observable
viability or non-orthodox seeds, such as many threatened and measurable factors and inputs appear to be the identical,
Magnolia species; the propagation from cuttings can also be crit- culture base qualities can slowly shift over time, whether by
ical for obtaining disease-free plants which limit the distribution to genetic drift or epigenetic adjustments, accumulation of micro-
other ex situ collections. Although tissue culture methods exist for scopic components of the medium, subtle effects of microcli-
some magnolia hybrids, effective micropropagation systems are mate, etc. Literally millions of micropropagated plants have
needed for wild magnolia species. The methodology includes col- been lost to disastrous events at some point in their production
lection from ex vitro, initiation, establishment, multiplication, and process. Infestations with insects, overnight meltdowns due to
rooting under in vitro conditions and hardening to ex vitro. endophytes, temperature shocks from AC outages. The primary
Initiation and establishment may be critical depending on the spe- normal losses are during transitioning, from open stomata and
cies, state, and stage of the explant. One of the main issues to reduced epidermal waxes leading to rapid desiccation, from
propagate magnolia shoots is due to the oxidation of phenolic bacterial endophytes (regardless of pathogenicity), from eco-
compounds during micropropagation (JunLi, 2007), and the way nomics: cost downturns, shifts and trends in marketplace.
to overcome this problem varies inside of the genus. An in vitro More subtle losses can occur as the result of air stagnation in
repository was created at the Huntington Library, Art Museum and closed rooms, water purification systems becoming contami-
Botanical Gardens (San Marino, CA) for the ex situ conservation nated with microbial endotoxins, vessel differences in gas ex-
of magnolia species, thus reducing the maintenance cost. Shoots of change and humidity. It would be useful if the known pitfalls
different Magnolia spp were introduced in tissue culture, and sev- facing the scale-up of micropropagated plants could be antici-
eral trials have been performed to study the effect of the media pated and prevented, but this is not always possible.
composition on the establishment, multiplication, and rooting. The Consequently, strategies are still needed for intervention and
hardening method has been optimized to reduce loss during the recovery once a problem is observed. Strategies for 1) manage-
acclimation from in vitro to ex vitro conditions. Besides, cryopres- ment of biological contaminants, such as bacteria, common
ervation experiments resulted in the first successful method to thrips, and fungi, as well as the notoriously disastrous mites,
cryopreserve magnolia shoot tips (Folgado and Panis, 2018), will be presented. Recommended quality assurance and control
which will allow the long-term conservation of those trees. protocols will be outlined.PLANT SYMPOSIA AND WORKSHOPS
P-3 OF OVEREXPRESSION OF CONSTANS 1 gene enhanced berry
productivity through the interaction of MIKC MADS-box genes.
Overview of Plant Tissue Culture Media and Practices. Overexpression of a blueberry DWARF AND DELAYED
GREGORY C. PHILLIPS. Arkansas State University, FLOWERING 1 (VcDDF1) increased freezing tolerance without
Arkansas Biosciences Institute, University of Arkansas System a trade-off impact on yield. Turning on a blueberry a RESPONSE
Division of Agriculture, PO Box 1080, State University, AR REGULATOR 2-like gene (VcRR2) in a mutant caused by a blue-
72467–1080. Email: gphillips@astate.edu berry DWARF AND DELAYED FLOWERING 1 (VcDDF1)
transgene insertion resulted in reduced chilling requirement for
This session is focused on best practices for plant tissue cul- flowering and a high yield potential. We also investigate blue-
ture. The choice of basal medium is critical for optimal re- berry FLOWERING LOCUS D, TERMINAL FLOWER 1 and
sponse, and may vary according to the species and purpose an apple FLOWERING LOCUS C gene in transgenic blue-
of the culture. This presentation will review the most com- berries. Overall, we use blueberry as a model to reveal the
monly used basal media as documented in recent literature, flowering mechanism in woody plants. We have demonstrated
and discuss considerations for further optimization or changes that manipulation of flowering pathway gene(s) or hormone
during culture. Screening of and manipulation of plant growth synthesis pathway gene(s) is a powerful approach to increase
regulators also is critical for achieving many developmental fruit productivity.
responses in culture. This presentation will review the most
commonly used plant growth regulators as documented in
recent literature, and discuss considerations for their manipu-
lation to achieve desired goals. Finally, the classical models P-5
for phytohormone manipulation for achieving organogenesis
or somatic embryohgenesis will be reviewed along with elab- Rapid Cycle Breeding of Tree Species. ANN CALLAHAN,
orated versions of these models as guides for the fundamental Ralph Scorza, and Chris Dardick. USDA-ARS Appalachian
practice of plant tissue culture. Fruit Research Station, 2217 Wiltshire Road, Kearneysville,
WV 25430. Email: Ann.Callahan@usda.gov
P-4 The genetic improvement of tree species has been a perennial
challenge to the breeding community due in large part to the long
Flowering Mechanism in Woody Plants: Manipulation of generation cycles of most fruit, forest, and ornamental tree species.
Blueberry Flowering Pathway Genes for Accelerated Breeding Breeding to address consumer demands, disease and insect resis-
and Yield Increase. GUO-QING SONG. Plant Biotechnology tance, improved production characteristics, and more recently ad-
Resource and Outreach Center, Department of Horticulture, aptation to climate change must be accelerated in order to foster
Michigan State University, East Lansing, MI 48824. Email: the development of productive orchards and sustainable forest
songg@msu.edu systems. To meet these challenges the development and release
of new improved varieties is more vital than ever. Yet, tree breed-
Blueberries contain high amounts of antioxidants known to be ing remains a slow and costly process often taking multiple
important for human health. Developing new cultivars with dif- breeders’ lifetimes to produce trees with significant genetic im-
ferent chilling requirement, high cold/heat tolerance, and for high provements. Much research has focused on marker assisted selec-
yield are among the top priorities in blueberry breeding due par- tion, sequencing for candidate gene identification, and genetic
ticularly to the anticipation of climate changes and the rapidly engineering as means to advance tree fruit breeding. However,
expanding market need of blueberry products. We have devel- these strategies remain limited by inherently slow generation cy-
oped a very efficient protocol of Agrobacterium tumefaciens-me- cles. Transgenic plants overexpressing Flowering Locus T (FT)
diated transformation of blueberries. We use stable transgenic and other flowering regulatory genes have been shown to express
blueberries, transcriptomic analysis and phytohormone analysis significantly shorter juvenility periods, typically one year or
to reveal blueberry flowering mechanism. Transgenic blueberries less. Rapid cycle breeding is being utilized in apple and
overexpressing a blueberry FLOWERING LOCUS T (VcFT) fa- Prunus domestica plum improvement programs where intro-
cilitate FAST-TRACK blueberry breeding through either gression of traits is being accomplished in a few years rather
transgrafting or crossing. Transgrafting on VcFT-overexpressing than a few decades. While this breeding system leverages
blueberry plants promotes floral bud formation in nontransgenic genetically engineered plants to accelerate breeding, the out-
scions and thus not only demonstrates that hormones are involved put is conventional cultivars, as the ectopically expressed
in FT-induced long-distance transport of the florigenic signals but flowering gene can be selected against to yield non-
also provides a new approach to increase blueberry yield. transgenic progeny that in the US do not require regulatory
Overexpression of the K-domain of a blueberry SUPPRESSOR approval.PLANT SYMPOSIA AND WORKSHOPS
P-6 Jolla, CA, 92037; and 4Howard Hughes Medical Institute,
Salk Institute for Biological Studies, La Jolla, CA, 92037.
Deciphering the Molecular and Cellular Development of a C4 Email: wbusch@salk.edu
Photosynthetic Leaf. TAMMY L. SAGE. University of
Toronto, Department of Ecology & Evolutionary Biology, Human activities have resulted in a continuous increase of CO2 in
25 Willcocks Street, Toronto, ON M5R3C6, CANADA. the earth’s atmosphere. This increase is already causing drastic
Email: tammy.sage@utoronto.ca alterations of the earth’s climate and will continue to change it
drastically. Despite efforts to reduce them, human caused carbon
Global agriculture has reached a time of crisis. Population emissions have increased over the past years. Carbon drawdown
growth and climate change are causing food shortages that technologies are needed to mitigate human CO2 emissions during
are undermining global food security. Equally as significant, the transition to renewable energy and then beyond to remove
increases in crop yields facilitated by the “Green Revolution” CO2 from the atmosphere. Plants are key components of the
are approaching their maximum potential resulting in a slowing global carbon cycle. Their ability to fix atmospheric CO2 exceeds
of agricultural productivity. To this end, there has been a call for human emissions by many times, but the vast majority of the plant
a second “Green Revolution” enabled by novel technologies to materials is degraded quickly. Soils are a major sink for stable
raise the ceiling on crop productivity in a warming climate. plant derived carbon and soil contains more than three times the
Increasing the yield ceiling is best done by improving photo- amount of carbon that is present in the atmosphere. Plants roots
synthetic efficiency. The most important crop plants use the C3 are a major source of soil carbon. Not only are high levels of soil
photosynthetic pathway wherein the efficiency of photosynthe- carbon usually associated with improved soil health but deposit-
sis is inhibited by the wasteful process of photorespiration, ing increased amounts of soil carbon from plants will drawdown
particularly at warmer temperatures. C4 photosynthesis in crops atmospheric CO2. We have identified several root traits that prom-
such as maize enhances yields by up to 50% over that of C3 ise to be key to enhance carbon deposition and carbon residence
plants by inhibiting photorespiration. Engineering the C4 path- time in soil. We are using rapid prototyping strategies in model
way into C3 crops is one way to increase the yield ceiling and plans to identify key genes and pathways to improve these traits in
forestall hunger in the twenty-first century. The spatial separa- several crop species.
tion of the initial fixation of atmospheric carbon by phospho-
enolpyruvate carboxylase from the secondary refixation of CO2
by RuBP carboxylase/oxygenase is an essential feature of the P-8
C4 carbon concentrating mechanism. This architectural rela-
tionship is present in roughly 8100 C4 species regardless of Engineering the Nitrogen-fixing Nodulation Trait
whether reactions are occurring within single-cells or more Using Legumes and the Tropical Parasponia Tree
commonly, between two-cell types, mesophyll (M) and a (Cannabaceae) as Templates. R. GEURTS. Wageningen
sheath of cells surrounding vascular tissue. Evidence from lin- University, Department of Plant Science, Laboratory of
eages possessing two-celled C4 photosynthesis supports a grad- Molecular Biology, Wageningen, THE NETHERLANDS.
ual model of C4 origins from C3 ancestors with early phases Email: rene.geurts@wur.nl
classified as C2 photosynthesis (photorespiratory glycine shut-
tle). Photosynthetic activation of sheath cells, modifications in Nitrogen-fixing root nodules are a common trait of legumes
cell division and expansion patterns of sheath and M cells as (Fabaceae, Fabales). These nodules are the result of a mutualistic
well as increase plasmodesmata density between sheath and M endosymbiosis between the plant root and diazotrophic rhizobium
cells underpin the evolution of the two-celled CCM. Key mo- bacteria. Besides legumes, nitrogen-fixing root nodules can be
lecular and cellular signatures important for regulating C4 leaf found in nine other taxonomic lineages in the related orders
development in both eudicots and monocots will be presented. Fagales, Rosales and Cucurbitales having either rhizobium or
Frankia as microsymbiont. Recent phylogenomic research sug-
gests that nodulation has a single evolutionary origin. A long-
P-7 standing research question is whether this nitrogen-fixing nodula-
tion trait can be engineered into crop plants. However, the key
Harnessing Plant Biology to Address Climate Change. W. genetic adaptations allowing the formation of nitrogen-fixing root
BUSCH1,2, J. Chory1,2,4, J. Law1,2, and J. P. Noel1,3,4. 1Salk nodules have not yet been identified. We use nodulating
Institute for Biological Studies, Harnessing Plants Initiative, Parasponia trees (Cannabaceae, Rosales) as a comparative sys-
La Jolla, CA 92037; 2Salk Institute for Biological Studies, tem. By establishing in vitro propagation, Agrobacterium
Plant Molecular and Cellular Biology Laboratory, La Jolla, tumefaciens-mediated transformation and CRISPR-Cas9 muta-
CA 92037; 3Jack H. Skirball Center for Chemical Biology genesis in Parasponia, we aim to identify a genetic blueprint for
and Proteomics, Salk Institute for Biological Studies, La engineering the nitrogen-fixing nodulation trait.PLANT SYMPOSIA AND WORKSHOPS
P-9 Medicinal and recreational Cannabis is most often vegetative-
ly propagated from mother plants to maintain genetic and
Ignorance is Bliss? An Overview of Diseases Affecting phenotypic uniformity. However, long term maintenance of
Cannabis. D. L. JOLY, C. Balthazar, V. Blanchet, C. genetics as mother plants requires significant amounts of valu-
Cormier, A. J. Cull, N. Pépin, and F. O. Hébert. Université able space and leaves them vulnerable to insect, disease, and
de Moncton, 18 avenue Antonine-Maillet, Moncton, NB E1A viral infections that are then transmitted to the propagules.
3E9, CANADA. Email: david.joly@umoncton.ca Given the lack of registered pest control products for
Cannabis, it is critical to start with clean, healthy planting
Supplying cannabis and its derivatives to a legalized retail material. Plant tissue culture and micropropagation provide
market represents a major economic opportunity. Like any the foundation for the maintenance of genetics free from such
other crop, cannabis is susceptible to various diseases. threats and facilitates the production of clean planting material
Mitigating crop losses caused by these diseases offers tremen- for commercial production. However, due to the long history
dous opportunities for increased production, ensuring a con- of prohibition, very little research has been published for this
tinuous access to cannabis. Pathogens can directly impact crop and current research is limited to very few genotypes.
growth by affecting roots, crown or foliage. These pathogens Early experience in our lab demonstrated that published pro-
can also colonize inflorescences during not only development, tocols are not reproducible across most genetics and highlight-
but also following harvest, thereby reducing product quality. ed the need for further development. This talk will provide an
Beyond concerns of product quality, recent legislation has led overview of our recent work on in vitro techniques for
to mandatory testing for the presence of certain microbes pos- Cannabis including aspects of media optimization, various
ing a threat to human health, adding a consumer safety barrier non-traditional approaches to micropropagation, the effects
that will lead to more lost product. Because production of of plant growth regulators, photoautotrophic culture systems,
cannabis has been illegal for decades, it has not benefitted and other aspects of our research. Overall, this presentation
from the application of biotechnologies toward the develop- will highlight a number of advances in Cannabis tissue culture
ment of disease-resistant varieties, which have led to increases to help develop more efficient methods for researchers and
in yields and sustainability in other crops. Furthermore, the industry.
number of pesticides registered for use on cannabis is current-
ly limited. In this context, the use of in vitro biology for the
production of pathogen-free plant materials shows promise in
avoiding the propagation of diseases when subculturing from P-11
mother plants. This presentation reviews state of knowledge
and challenges in cannabis pathology, and highlights future In Vitro Technologies for Clean Plant Production in Cannabis.
research avenues to improve cannabis cultivation and allow JEREMY WARREN. Dark Heart Nursery, 717 Kevin Ct.,
the harvesting of high-quality products. A project aiming at Oakland, CA 94621. Email: jwarren@darkheartnursery.com
referencing diseases affecting cannabis will be described.
Through this project, a DNA barcode reference library for Traditional commercial scale agronomic crops have benefited
pathogens is being established, where georeferenced records from decades of governmental and university-backed clean
and their corresponding DNA sequences are integrated in a plant programs, which provide certified pathogen-free plant-
Barcode of Life Data Systems database. This resource will ing stock for many agronomically important crops. These cer-
allow the development of DNA-based detection tools, crucial tified pathogen-free plants often serve as foundational plant-
for the monitoring of pathogens, the identification of their ing materials for agricultural companies, as having pathogen
sources and pathways, and the prediction of outbreaks. free material can greatly enhance breeding timelines and lead
Culture collections are also maintained, a pivotal resource to to positive outcomes for growers’ crops. Until recently gov-
allow the identification of resistant varieties. ernment and university funding for Cannabis sativa research
has been minimal and mainly focused on human medicinal
uses. This has left a huge knowledge void on the plant sci-
ences side of C. sativa research, especially around understand-
P-10 ing which pathogens are important in C. sativa cultivation and
subsequently the development of a C. sativa clean plant pro-
Advances in Micropropagation of Cannabis. MAX JONES. gram. As Director of Plant Health at Dark Heart Nursery, Dr.
Gosling Research Institute for Plant Preservation, Department Jeremy Warren will discuss why developing a clean plant
of Plant Agriculture, University of Guelph, Guelph, ON, program to create certified pathogen-free stock is critically
CANADA. Email: amjones@uoguelph.ca important for the Cannabis industry and how to use in vitroPLANT SYMPOSIA AND WORKSHOPS
technologies to create cannabis stock that is free from patho- quality of image data from in vitro fractional factorial screening
gens, such as, Hop latent viroid, a recently described cannabis and response surface optimization experiments will be illustrated
pathogen causing major yield loses throughout the US and and the resulting analyses compared to the analyses using stan-
Canada. dard measures. The impact on experimental design when using
image data, that is often of higher consistency and resolution than
standard measures, will be discussed.
P-12
Maximizing the Value of Your Imaging Data with High- P-15
content Image Analysis and Deep Learning. BETH CIMINI,
Allen Goodman, Shantanu Singh, Juan Caicedo, and An Overview of Epigenetics in In Vitro Cultured Plants. C.
Anne Carpenter. Broad Institute of MIT and Harvard, DE-LA-PEÑA, F. Duarte-Aké, and R. Us-Camas. Centro de
415 Main Street, Cambridge, MA 02155. Email: I n v e s t i g a c i ó n C i e n t í fi c a de Yu c a t á n , U n i d a d d e
bcimini@broadinstitute.org Biotecnología, Calle 43 # 130 × 32 y 34 Chuburná de
Hidalgo C.P. 97205, Mérida, Yucatán, MÉXICO. Email:
While standard biological images contain a minimum of mil- clelia@cicy.mx
lions of pixels of quantitative information, in many cases the
richness of this information is not fully mined when determin- In vitro plant cell and tissue culture techniques are the basis of
ing phenotypes. High content image analysis tools like many micropropagation and breeding programs for scientific
CellProfiler can help users extract more information easily, research. Plant tissue culture involves organogenesis and em-
allowing biologists from many fields to generate many more bryogenesis, and the outcome depends on the different condi-
measurements of each cell or object than was previously pos- tions to which the tissue is exposed. New data has come out
sible; applying pretrained neural networks can create similarly about a connection between plant morphogenesis and epige-
rich measurement sets. Either type of measurement can then netics. Epigenetics is a very sensitive regulatory mechanism,
be used to perform morphological profiling, which allows which in most of cases is affected by the environment.
users to use this extra “hidden” information to make robust Although it is known that, under plant morphogenesis, the
connections between treatment groups in their experiments. genome has little or no change, DNA methylation and histone
We will describe new tools our lab is creating to leverage these modifications are very susceptible to those in vitro environ-
measurements, as well as broader goals for tool development mental conditions. In the present talk, we highlight the most
in image analysis for the 2020s. used in vitro systems such as organogenesis and somatic em-
bryogenesis in plants and discuss how epigenetics plays a
pivotal role in the phenotype outcome. Recent work using
P-13 an albino Agave phenotype will be also presented, in which
we suggest that an “epigenetic stress memory” during in vitro
In Vitro Data Collection Using Image Analysis and Machine conditions causes a chromatin shift that favors the generation
Learning. R. P. NIEDZ. U. S. Department of Agriculture, of variegated and albino shoots. Projects are supported by the
Agricultural Research Service, U. S. Horticultural Research Consejo Nacional de Ciencia y Tecnología (CONACYT).
Laboratory. Email: randall.niedz@usda.gov
Image analysis is a broad field and covers the collection of images P-17
and the processing of those images to extract data for information
on growth, quality, and diagnostics of disease and physiological Overcoming Obstacles Associated with Woody Plant
disorders. Image data is a useful measure for quantifying in vitro Micropropagation. MICAH E. STEVENS and Reid
culture systems because it is accurate, simple to collect, and reli- Robinson. Sierra Gold Nurseries, 5320 Garden Highway,
able. Incorporating image analyses into the routine collection of Yuba City, CA 95991. Email: Micah@sgtrees.com
data is possible because of the availability of free, inexpensive,
and user-friendly image analysis and machine learning software. Tissue culture propagation is ideal for quickly producing large
Some widely available, well-supported, and free software tools numbers of unique genotypes, circumventing problems with
will be described. Image analysis and machine learning of data, traditional vegetative propagation, maintaining genetic diver-
acquired using ordinary digital cameras, will be illustrated using sity, accelerating breeding programs, and conservation of en-
examples from citrus in vitro and ex vitro cultures and will include dangered or threatened species. While the principles of plant
embryogenic and nonembryogenic cell lines, micropropagated tissue culture have been established for decades the
shoot cultures, greenhouse plants, and field trees. The use and micropropagation of woody plants continues to encounterPLANT SYMPOSIA AND WORKSHOPS
many hurdles. These obstacles are exacerbated and amplified P-19
in commercial settings leading to increased production costs
for agronomically and horticulturally valuable genotypes. Recognition and Quantitative Analysis of Transformation
Endogenous, cryptic microbiota, and prolific phenolic exuda- Rate in Tissue Cultures Using Hyperspectral Imaging and
tion of some woody plants can quickly lead to explant necrosis Machine Learning. STEVEN H. STRAUSS 1 , Michael
without regular and frequent subculture to fresh medium dur- Nagle 1 , Jialin Yuan 2 , Damanpreet Kaur 2 , Jia Yi Li 2 ,
ing explant initiation. Sub- or supraoptimal concentrations of Cathleen Ma1, Ekaterina Peremyslova1, Yuan Jiang3, and
cytokinins used during the multiplication phase can lead to Fuxin Li2. 1Department of Forest Ecosystems & Society,
many deleterious effects, including stunted growth, vitrifica- Oregon State University, Corvallis, OR; 2Department of
tion, fasciation, shoot tip necrosis, and chlorosis have had Computer Science, Oregon State University, Corvallis, OR;
significant impacts on micropropagation success. Elevated and 3 Statistics Department, Oregon State University,
levels of phytohormones can persist in plant tissue after mul- Corvallis, OR. Email: Steve.Strauss@Oregonstate.Edu
tiplication and lead to unforeseen physiological abnormalities.
Finally, the interaction between genotype and basal salt or Phenotyping of in vitro plant transformation relies on systems
phytohormone type and concentration can significantly influ- for recognizing and quantifying transgenic tissues. Reporter
ence propagation success. High variability among species, proteins (e.g., GFP) are commonly used, however, their utility
even within in the same genus, to identical protocols contrib- is limited by the time-cost of screening and quantifying large
utes to many of the difficulties with woody plant tissue cul- cohorts of tissue cultures. This limitation is especially impor-
ture. While responses to in vitro environments are often geno- tant for large scale studies such as mutant screens, commercial
type specific, and are controlled by many endogenous and programs that seek to produce and screen dozens to hundreds
exogenous factors, tissue culture remains integral to produce of transgenic events, and genome-wide association studies
high-valued plants on a commercial scale. Taken together (GWAS) of in vitro expressed characteristics. To enable
these issues pose significant difficulties to woody plant GWAS of in vitro transformation in Populus trichocarpa,
micropropagation, however many of these can be mitigated we developed a phenomics workflow that provides measures
or circumvented with the right techniques. of reporter protein signal in specific in vitro tissues by cross-
referencing of results from separate analyses of RGB and
hyperspectral images: 1) Tissue cultures are imaged using a
custom instrument produced by Middleton Spectral Vision
P-18 (Wisconsin) that collects RGB images and hyperspectral im-
ages of fluorescent proteins excited by a blue laser. 2) Deep
Recent Developments in Imaging Analytics, Artificial learning is used to segment RGB images according to the
Intelligence and Robotics. BALATHASAN GIRITHARAN. stage and tissue type (e.g., callus, shoot) during regeneration.
Bayer Crop Science, 800 North Lindbergh Blvd., St. Louis, 3) Linear regression is then used for analysis of hyperspectral
MO 631671. Email: balathasan.giritharan@bayer.com images, providing a measure of fluorescent proteins for each
pixel of the image. By cross-referencing these complementary
With the significant advancement in machine learning, datasets, we obtain statistics for reporter protein signal in spe-
neural networks, and fast computational units, artificial cific tissues which can then be summed to estimate transfor-
intelligence is getting closer to enabling machines to mation rates for specific tissues and populations of explants.
think more like humans. Moreover, imaging sensors To demonstrate the effectiveness of this method for phenotyp-
have become cheaper, smaller, more reliable, and can ing in vitro transformation, we will compare the information
capture multiple modalities, including hyperspectral, x- content and precision of human-scored statistics to those pro-
ray, fluorescence, LiDAR. With sensors becoming ubiq- duced by our phenomics workflow. We thank the National
uitous, the amount of data generated and stored is grow- Science Foundation Plant Genome Research Program (IOS
ing exponentially. Sensors function as ears and eyes of # 1546900) as the primary source of financial support. We
AI, and the success of AI systems depends on data used also thank the GREAT TREES (Genetic Research on
to train the models used in the AI system. With a large Engineering and Advanced Transformation of Trees) consor-
amount of sensor data becoming available to help the tium at Oregon State University for partial support.
AI systems to get smarter. In this presentation, we dis-
cuss the landscape of techniques in artificial intelli-
gence, imaging, and robotics in the field of biology P-20
and how AI is to perform tasks that generally require
human knowledge, visual perception, and decision- RoBo®Cut a Revolutionary Invention for In Vitro Plant
making. Tissue Culturing, RoBoCut-smart Propagation, Better Plants.PLANT SYMPOSIA AND WORKSHOPS
STEPHEN VON RUNDSTEDT. Bock Bio Science GmbH, the target locus from the cellular response to DSBs.
Butendieker Landstr. 49A, 28357 Bremen, GERMANY. Recently, a new genome editing methodology called ba-
Email: max@breedingleaders.com se editing was developed that allows the precise, irre-
versible conversion of one base pair to another via the
In vitro Plant Tissue Culture (“PTC”) has been exposed to direct chemical modification of the DNA nucleobases.
high competitive pressure for years. As in many industrial This is accomplished through the engineering of fusions
fields the reduction of personnel costs as well as scaling up of CRISPR/Cas9 and ssDNA modifying enzymes. The
the production line are the ascendant factors nowadays. resulting fusions retain the ability to be programmed
Consequently, many companies have been forced to outsource with a guide RNA and do not induce DSB but rather
their productions to low-wage countries despite the immense mediate the direct conversion of cytosine to uracil
risks attached to it. As a leading plant breeding company in (thereby effecting a C•G to T•A substitution) or adeno-
Germany, the family-owned Bock Bio Science GmbH strong- sine to inosine (effecting an A•T to G•C substitution).
ly believes that healthy sustainability is only achievable with Here I will describe our experimental and computational
regional business growth without exploitation of geographical investigations to understand the enzymatic mechanism
profits. To make this possible, we will introduce our revolu- of adenine deamination by the adenosine base editor. I
tionary invention called “RoBo®Cut” in this session. will additionally discuss our efforts to understand the
RoBo®Cut is a fully autonomous, automated in vitro plant cellular processing of U•G lesions, the intermediate of
propagation system that is able to provide the most relevant cytosine base editing.
manufacturing step of “separating plants” in plant propaga-
tion. With the combination of several robot arms, 3D vision,
Artificial Intelligence and a laser system, it is able to precisely P-22
cut the plantlets along their natural growth lines without hu-
man interaction. Via smart perspective analysis RoBo®Cut Homologous and Non-homologous Somatic Recombination
already succeeds in placing the cut plantlets into a fresh me- Leads to Genome Remodeling During Clonal Growth of
dium every 6 seconds, whereby the entire chain of capturing a Potato. L. COMAI1, K. R. Amundson1, B. Ordoñez1, X.
donor plant, creating the dividing line and cutting via laser is Zhao1, G. T. Braz2, J. Jiang2, and Isabelle M. Henry1.
1
run through in advance. In addition, RoBo®Cut’s unique de- Department of Plant Biology and Genome Center, University
sign allows to propagate all kind of plants within the Plant of California Davis, Davis, CA and 2Department of Plant
Tissue Culture phase while guaranteeing highest sterility. Biology, East Lansing, MI. Email: lcomai@ucdavis.edu
With the help of this invention, we are already able to reduce
manufacturing costs dramatically today whilst raising the Genome integrity is dependent on controlling and
overall plant quality and performance of the plants. Thus, balancing DNA replication, recombination, cell division,
we believe RoBo®Cut will bring PTC back onshore to the mutation, and transposition. The orchestration of these
high wage countries and provide an affordable technology for processes can fail resulting in genome instability, an
many crops that as of today cannot be made via PTC because outcome well documented during stress such as when
of costs, while helping us to reduce the transportation as well cultured cells are induced to regenerate plants. The
as the greenhouse emissions to a minimum. resulting “somaclonal variation” appears extreme, but
similar processes may occur less frequently during nor-
mal growth. Plants’ flexible ploidy, meristematic
P-21 growth, and lack of predetermined germline might en-
able retention of karyotypic novelty, perhaps explaining
Investigating the Chemical and Cellular Mechanisms of Base the formation of sports during clonal growth. We ex-
Editing. A. C. KOMOR. Department of Chemistry and plored this hypothesis using potato, in which we docu-
Biochemistry, University of California, San Diego, La Jolla, ment non-homologous and homologous recombination
CA 92093. Email: akomor@ucsd.edu preferentially located on certain chromosomes. These
events are consistent with fragile sites and can result,
Most genome-editing technologies introduce double- alternatively, in chromoanagenesis or copy-neutral loss
stranded DNA breaks (DSBs) at a target locus as the of heterozygosity. Our observations provide a frame-
first step to gene correction. These methods are typical- work for understanding both genome instability and its
ly inefficient at point mutation correction and induce an consequences. In addition, they suggest strategies for
abundance of random insertions and deletions (indels) at engineering of plant genomes.You can also read
