is essential to know the role of specific small RNA followed by genetic manipulation
for improvement of drought stress tolerance in soybean crop. The RNAi technology
has been successfully used for improvement of soybean crop in terms of enhancing
abiotic stress tolerance (Wang et al. 2015; Srivastava et al. 2017; Li et al. 2017a;
Mao et al. 2018). RNAi has been effectively utilized for incorporating desired traits
for abiotic stress tolerance in various plant species (Jagtap et al. 2011; Pradhan et al.
2015; Meena et al. 2017; Li et al. 2017a). Wang et al. (2015) studied the interaction
of GmWRKY27 with GmMYB174 and reported that these two cooperatively inhibit
transcription of GmNAC29 by binding to the core sequences in its promoter. The
downregulation of expression of GmNAC29 leads to reduced intracellular ROS
levels. The GmWRKY27 may also increase proline content by indirectly suppressing
the transcription of PDH which ultimately led to improvement in stress tolerance in
soybean (Wang et al. 2015). Li constructed soybean GmRACK1 silenced (RNA
interference, RNAi) and over-expressing plants. The GmRACK1-RNAi lines
showed significantly improved drought stress tolerance while the over-expressing
seedlings were hypersensitive to drought stress when compared to wild-type in terms
of plant survival rates after 10 days of drought. GmRACK1-RNAi plants were found
to be more sensitive to ABA when seeds germinated and root grew.
4.9.3 Genome Editing Based Techniques
The availability of soybean wild species and genetic variations in soybean germplasm is crucial for soybean improvement programs targeting drought toleance.
However, the lack of enough natural germplasm, genetic diversity, and mutant
collections limits both basic and applied research, particularly in soybean. The
genome editing tools provide opportunity to overcome these limitations via creation
of such variations in the genome of crop plants. Such approaches can reduce
breeding or gene transformation time greatly for production of new varieties/transgenic plants with desired traits, such as abiotic stress tolerance. The CRISPR
technology is being seen as an advancement of plant breeding technologies.
Non-transgenic approaches are also available for delivery of such nucleases to
produce mutant plants (Marton et al. 2010). As a result, crop varieties produced
using these technologies may qualify as non-GM and would have enormous impact
on plant biotechnology and breeding. There are four genome editing tools,
meganucleases, zinc-finger nucleases (ZFN), Transcription Activator-like Effector
Nucleases (TALEN) and the Clustered Regularly Interspaced Short Palindromic
Repeat (CRISPR)/CRISPR-ASsociated nuclease protein (Cas) system, which have
provided targeted gene modification in plants (Cermak et al. 2015; Gao et al. 2010;
Li et al. 2012, 2013a, b; Shukla et al. 2009). Among these, the CRISPR-Cas9 system
is the easiest to implement and is highly efficient. The system consists of a Cas9
endonuclease derived from Streptococcus pyogenes and a chimeric single guide
RNA that directs Cas9 to a target DNA sequence in the genome. The CRISPR-Cas9
genome editing is accomplished by introducing a DNA double-strand break in the
target locus by nuclease enzyme named Cas9, followed by DNA repair through
4 Breeding and Molecular Approaches for Evolving Drought-Tolerant Soybeans
111
for improvement of drought stress tolerance in soybean crop. The RNAi technology
has been successfully used for improvement of soybean crop in terms of enhancing
abiotic stress tolerance (Wang et al. 2015; Srivastava et al. 2017; Li et al. 2017a;
Mao et al. 2018). RNAi has been effectively utilized for incorporating desired traits
for abiotic stress tolerance in various plant species (Jagtap et al. 2011; Pradhan et al.
2015; Meena et al. 2017; Li et al. 2017a). Wang et al. (2015) studied the interaction
of GmWRKY27 with GmMYB174 and reported that these two cooperatively inhibit
transcription of GmNAC29 by binding to the core sequences in its promoter. The
downregulation of expression of GmNAC29 leads to reduced intracellular ROS
levels. The GmWRKY27 may also increase proline content by indirectly suppressing
the transcription of PDH which ultimately led to improvement in stress tolerance in
soybean (Wang et al. 2015). Li constructed soybean GmRACK1 silenced (RNA
interference, RNAi) and over-expressing plants. The GmRACK1-RNAi lines
showed significantly improved drought stress tolerance while the over-expressing
seedlings were hypersensitive to drought stress when compared to wild-type in terms
of plant survival rates after 10 days of drought. GmRACK1-RNAi plants were found
to be more sensitive to ABA when seeds germinated and root grew.
4.9.3 Genome Editing Based Techniques
The availability of soybean wild species and genetic variations in soybean germplasm is crucial for soybean improvement programs targeting drought toleance.
However, the lack of enough natural germplasm, genetic diversity, and mutant
collections limits both basic and applied research, particularly in soybean. The
genome editing tools provide opportunity to overcome these limitations via creation
of such variations in the genome of crop plants. Such approaches can reduce
breeding or gene transformation time greatly for production of new varieties/transgenic plants with desired traits, such as abiotic stress tolerance. The CRISPR
technology is being seen as an advancement of plant breeding technologies.
Non-transgenic approaches are also available for delivery of such nucleases to
produce mutant plants (Marton et al. 2010). As a result, crop varieties produced
using these technologies may qualify as non-GM and would have enormous impact
on plant biotechnology and breeding. There are four genome editing tools,
meganucleases, zinc-finger nucleases (ZFN), Transcription Activator-like Effector
Nucleases (TALEN) and the Clustered Regularly Interspaced Short Palindromic
Repeat (CRISPR)/CRISPR-ASsociated nuclease protein (Cas) system, which have
provided targeted gene modification in plants (Cermak et al. 2015; Gao et al. 2010;
Li et al. 2012, 2013a, b; Shukla et al. 2009). Among these, the CRISPR-Cas9 system
is the easiest to implement and is highly efficient. The system consists of a Cas9
endonuclease derived from Streptococcus pyogenes and a chimeric single guide
RNA that directs Cas9 to a target DNA sequence in the genome. The CRISPR-Cas9
genome editing is accomplished by introducing a DNA double-strand break in the
target locus by nuclease enzyme named Cas9, followed by DNA repair through
4 Breeding and Molecular Approaches for Evolving Drought-Tolerant Soybeans
111