This program targets the implementation of the tools of biotechnology as a means to introduce
novel input and output traits into soybean. The derived transgenic events expressing the
targeted trait(s) are characterized at the genetic level and subsequently phenotyped under
greenhouse conditions. Lead transgenic events are then selected based on the data gathered
from these initial genetic and phenotypic evaluations. Selected lead events per transgenic trait
are then characterized under field environments at UNL’s Plant Biotechnology Field Facility
for agronomics and stability of the trait(s) of interest. The traits currently being researched and
developed under this program include, strategies to boost photosynthetic capacity as a means
to enhance yield and/or protect yield under stress conditions, in both standard leaf and
lanceolate leaf genotypes of soybean. Moreover, through resources provided by USB these
photosynthetic transgenes are being combined with transgenes to elevate oil percentage,
without comprising protein quality or quantity. The second trait being targeted is alteration of
stomata (gas/water pores) numbers on the leaf surface, as a means to influence water use
efficiency in soybean. More recently the program is designing genetic approaches to produce a
soybean- based feedstock for plant-based burger formulations. Here genetic stacks are being
created and tested that combine high solids oil (high in stearic acid and oleic acid), coupled
with the synthesis of leghemoglobin in the seed. Lastly, the program is continuing to evaluate
soybean events carrying interfering RNAs designed to impede aphid feeding and developing
soybean lineages that harbor genome edited null mutations in four seed storage proteins as a
means to improve the quality of the protein reserve in the seed.

1) Design genetic strategies to boost carbon capture (photosynthesis) and 'pull' carbon towards oil during seed
development
2) Design genetic strategies to alter stomate number on the leaf surface to improve water use efficiency in soybean
3) Design a soybean that has valuable attributes for "veggie burger" formulations
4) Utilize genome editing tools for targeted improvement on protein quality and functionality in soybean
5) Continue to collaborate with colleagues for novel resistance approaches towards aphids and development of a
soybean-based feedstock for aquaculture

The photosynthetic component of the program will include continuation of the field trials with
selected lead events that carry the transgenic alleles shown to increase photosynthesis, in both
standard leaf and lanceolate leaf genetic backgrounds, under both irrigated and non-irrigated
environments. Moreover, the photosynthesis genes are being stacked with genes to increase
carbon flux to oil during seed development, to investigate if this approach will translate to a
soybean with higher oil, without impacting protein level. The data gathered over the past three
years on this set of novel soybean events is informing the next set of genetic designs to further
boost carbon capture and flux during seed development towards oil without compromising
protein level. This program will also characterize novel transgenic soybean events that have
reduced number of stomata on their leaves for drought tolerance. Here transgenic events are
being phenotyped under greenhouse conditions for alterations in stomata number. Those events
in which significant changes in stomata counts are observed will subsequently be evaluated
under field conditions. In addition, this program developed a single genetic element strategy that
leads to the production of a soybean oil with saturated fatty acids, approximately, 25% coupled
with oleic acid over 55%. Such an oil is a semisolid at room temperature and has uses in baking
applications, and other food formulations that require a solid oil for functionality. Support of this
program will enable us to continue field trials for more quantities and the subsequent evaluation
of the harvest for end use applications, in addition to the continuation of field testing of the
various input traits being developed in this program, along with monitoring influence of the
trait(s) on agronomic parameters.
Reagents have been assembled to create null mutations in four seed storage proteins (ßconglycinin,
glycinin, SAM22 and P34). These reagents have been introduced into soybean.
Genotyping and subsequent monitoring of the protein quantity/quality of the derived soybean is
ongoing. This project has built a genetic construct designed to synthesize leghemoglobin (key
ingredient to mimic texture/appearance of real beef in a veggie burger) in the seeds of soybean.
Soybean events have been generated that accumulate leghemoglobin in the seed at respectable
levels, however, the expected red coloration is only slightly observed. The lack of sufficient red
coloration can be attributed to multiple reasons, testing of two the possibilities is currently
ongoing.

Continued support of this program will enable the research and development of novel genetic
strategies that target means to influence quality and quantity of yield and/or protection of yield.
Outcomes from these studies hold great potential to increase productivity, and expand the soybean
market through improved oil functionality and protein quality. Importantly, this program is inherently
interdisciplinary tapping upon various scientific disciplines to address the various objectives or the
research activities.