**5. Next generation sequencing (NGS) and transcriptomics studies in okra**

The next-generation sequencing (NGS) technology has transformed the field of molecular breeding, particularly in the identification and development of SSR markers. The advantage of NGS techniques are cost efficiency and large number of SSR can be identified in shorter time [34]. There is limited literature available in okra related to studies using genomic approaches. Transcriptome analysis has appeared as a potential approach to identify the transcript/gene sequences in the crops like okra where limited or no genome sequence information is available. The first study on transcriptome assembly in okra was reported by [3]. Both leaf and pod tissues of okra were taken for RNA sequencing and short read assembly SRA accession no. SRX206126. They have identified more than 150,000 unigenes and 935 SSRs from unigenes (**Table 2**). These SSRs were used to study genetic diversity

**89**

**Okra Species** (*Abelmoschus* 

leaf and

Transcriptome

Illumina

26,324,557

150,000

935 SSRs

321 bp

SRX206126

[3]

unigenes

263

HiSeq™ 2000

assembly

pod

*esculentus* (L.)

Moench) CV. Mahnco

Arka Abhhay

*Abelmoschus esculentus*

Roots,

Transcriptome

Illumina HiSeq

716,330,252

293,971

—

1885 bp

SRP130180

unigenes

716

X Ten platform

assembly

stems,

and leaves

cv. Xianzhi

*Abelmoschus esculentus*

*Abelmoschus esculentus*

Leaves

Genome

Roche 454 GS

61,722

3735

402 SSRs

markers

contigs

FLX Titanium

assembly

cv. Arka Anamika

*Abelmoschus esculentus*

**Table 2.**

*Description of transcriptomic and genomic studies published in okra.*

Leaves

miRNA

Illumina and

207,285,863

845 novel

miRNAs

Ion torrent

identification

Leaves

Transcriptome

Illumina

206.3

66,382

9,578 SSRs

1,408 bp

SRX2995608,

[36]

SRX2995609,

SRX2995611, and

SRX2995612

—

PRJNA352593

[38]

[37]

unigenes

million

NextSeq 500

assembly

**Plant organ**

**Objective of study**

**Sequencing platform**

**Raw reads (M)**

**Final assembly**

**Marker discovery**

**N50**

**NCBI accession**

**Reference**

*Genomic Tools to Accelerate Improvement in Okra (*Abelmoschus esculentus*)*

[35]

*DOI: http://dx.doi.org/10.5772/intechopen.97005*


*Genomic Tools to Accelerate Improvement in Okra (*Abelmoschus esculentus*) DOI: http://dx.doi.org/10.5772/intechopen.97005*

> **Table 2.**

*Description of transcriptomic and genomic studies published in okra.*

*Landraces - Traditional Variety and Natural Breed*

**accessions/ genotypes**

*Abelmoschus escullentus* 66 (iPBS)-

**Type of markers No. of** 

*Abelmoschus escullentus* 48 ISSRS — 54.55% [24] *Abelmoschus escullentus* 24 ISSRS 22 0.531929 [19]

> retrotransposons and SSRs

*Abelmoschus spp* 39 RAPD 31 — [27] *Abelmoschus esculentus* 50 AFLP 33 12 [28] *Abelmoschus esculentus* 22 AFLP 8 0.26 [29] *Abelmoschus escullentus* 23 SRAP 39 — [22]

**primers**

83 iPBS, 9 SSRs

93 RAPD 13 — [26]

65 SSR 19 0.49 [3]

24 SSR 18 0.53 [30]

96 SSR 40 0.52 [31]

**PIC Reference**

[25]

0.66 iPBS 0.62 SSRs

**Species No. of** 

*Abelmoschus caillei* (50), *A. esculentus* 

*Abelmoschus esculentus*, *A. moschatus*, *A. manihot*

*Abelmoschus esculentus*

*A. tuberculatus* (1), *A. moschatus*(1), *A. manihot* (1)

*A. esculentus* (92) *A. tuberculatus* (1), *A. moschatus*(1), *A. moschatus subspecies tuberosus* (1), and *A. manihot*(1)

*Gene diversity studies in okra.*

*(43)*

(21)

**Table 1.**

obtained from assembled reads, a total of 2708 contigs had microsatellites. Finally 402 microsatellites were used for selection of 50 SSR primers for amplification in okra. This is the first report on the development of genomic SSR markers in okra

**5. Next generation sequencing (NGS) and transcriptomics studies** 

The next-generation sequencing (NGS) technology has transformed the field of molecular breeding, particularly in the identification and development of SSR markers. The advantage of NGS techniques are cost efficiency and large number of SSR can be identified in shorter time [34]. There is limited literature available in okra related to studies using genomic approaches. Transcriptome analysis has appeared as a potential approach to identify the transcript/gene sequences in the crops like okra where limited or no genome sequence information is available. The first study on transcriptome assembly in okra was reported by [3]. Both leaf and pod tissues of okra were taken for RNA sequencing and short read assembly SRA accession no. SRX206126. They have identified more than 150,000 unigenes and 935 SSRs from unigenes (**Table 2**). These SSRs were used to study genetic diversity

using next-generation sequencing technology.

**88**

**in okra**

in diverse okra germplasm by many workers and found informative for classification and understanding of okra germplasm. Ravishankar et al. [34] first reported development of genomic SSR markers in okra using Roche 454 Titanium pyrosequencing technology. A total of 61,722 reads were generated from 979,806 bp data. These reads were assembled into 3735 contigs of which 2708 had microsatellites. Primers were designed for 402 microsatellites, from which 50 randomly selected SSR primers were standardized for amplification of okra DNA.

MicroRNAs (miRNAs) are regulatory RNAs which plays a crucial role in regulating gene expressions at post-transcriptional levels in disease conditions. Vimala Kumar et al. [38] applied next generation sequencing approach for global profiling to characterize the miRNAs and their associated pre-miRNAs. Data analysis using miRPlant revealed 128 known and 845 novel miRNA candidates. They identified 57 known miRNAs of 15 families and 18 novel miRNAs. A total of 845 novel candidates were predicted when using cotton as a reference genome which is closely related to *A. esculentus.* In 2018, Zhang and co-workers used transcriptome approach to identify the transcripts involved in the synthesis of bioactive compounds like flavonoids and polysaccharides in various organs like roots, stems, leaves, flowers, and fruits. They have identified 293,971 unique unigene sequences, 931 unigenes related to enzymes of flavonoids biosynthesis were identified and quantified. 691 unigenes encoded 13 key enzymes related to fructose and mannose metabolism. The transcriptome data will be useful for the gene expression analysis study of the genes encoding bioactive compounds in okra. Priyavathi et al. [36] reported high quality leaf transcriptome of *A. esculentus* from leaf samples. 16,307 unigenes, 76 transcription factor, 9,578 potential SSRs have been identified from *A. esculentus* leaf transcriptome. The *A. esculentus* sequence information presented in this study will be a valuable resource for further molecular genetics and functional genomics studies for the improvement of this crop plant.

#### **6. Proteomic studies in okra**

Proteomics analysis is a tool to facilitate the study of global protein expression, and to provide a wealth of information on the role of individual proteins in specific biological processes. Due to the complex allopolyploid genome of okra little attention has been paid to the genetic improvement of this crop until recently. Soil salinity is one of the main abiotic stresses limiting plant growth and agricultural productivity. Understanding the mechanisms that protect plants from salt stress will help in the development of salt-stress-tolerant crop. Using TMT-based proteomic technique in 2019, Zhan and associates analyzed the differentially expressed proteins between the NaCl-treated seedlings and control. They have identified a total of 7179 proteins, there were 317 differentially expressed proteins (DEPs), of which 165 proteins were upregulated and 152 proteins downregulated in the presence of NaCl.

#### **7. Discussion**

The molecular markers can be effectively used to enhance okra breeding programme through marker-assisted selection (MAS). Marker assisted breeding allows selection of desired trait at early stage which leads to accelerated development of improved varieties. Although, molecular markers have been broadly employed for DNA fingerprinting, gene mapping and gene tagging, seed purity testing and to know the molecular basis of heterosis in various crops, but in okra its use is still

**91**

**Author details**

**8. Conclusion**

New Delhi, India

Suman Lata\*, Ramesh Kumar Yadav and B.S. Tomar

metabolic pathway governing disease resistance.

\*Address all correspondence to: sumanlata3@gmail.com

provided the original work is properly cited.

Division of Vegetable Science, ICAR-Indian Agricultural Research Institute,

© 2021 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/ by/3.0), which permits unrestricted use, distribution, and reproduction in any medium,

*Genomic Tools to Accelerate Improvement in Okra (*Abelmoschus esculentus*)*

limited, therefore, it is the need of hour to use these approaches to accelerate the okra breeding programme at faster pace. The genomics and bioinformatics should also be well integrated into the programme for effective application of markers to okra breeding. A comparative genomics approach of other crop should also be applied for breeding programmes of those crops where the genome information is not available. Development of cost-effective genotyping technologies should always be the integral part of any improvement programme. There is need to use SSR and SNP based genotyping technologies as well as advanced technologies such as next

Resilient resistance to begomoviruses like Yellow vein mosaic virus (YVMV)

Okra is considered as a non model crop with a complex genome. Genomic studies like genome sequencing and transcriptome sequencing will help in identification of genes/transcripts for important agronomic traits like disease resistance in okra. Tools like RNAi and CRISPR/Cas9 genome editing can be employed for imparting resistance as well as functional characterization of genes. Identification of genes/ transcripts and markers linked to the resistance genes will help in breeding for resistance varieties. Also, there is requirement to bred durable/stable resistance against multiple diseases. ELCV is emerging as new havoc for okra along with YVMV which may be more difficult for production of okra in future. Therefore, gene pyramiding for combating multiple disease resistance genes in various genetic backgrounds should be done. There is also need to breed varieties/hybrids tolerant to abiotic stresses like cold, moisture and salt stress in the changing climatic scenario.

poses a serious challenge to both breeders and pathologists as these viruses are highly diverse, and constantly generate new forms via recombination. Biotechnological tools to generate resistant cultivar against Yellow vein mosaic disease (YVMD) are limited due to the lack of informative polymorphic markers, genetic maps and genome sequence information. Therefore, use of novel molecular and genomic tools will help in the accomplishing resistance against YVMV in okra. Identification of markers linked to the YVMV resistance gene/s and its pyramiding for combining multiple disease resistance genes in various backgrounds will help in okra crop improvement. In addition, genomic tools will help in elucidating the

*DOI: http://dx.doi.org/10.5772/intechopen.97005*

generation sequencing.

#### *Genomic Tools to Accelerate Improvement in Okra (*Abelmoschus esculentus*) DOI: http://dx.doi.org/10.5772/intechopen.97005*

limited, therefore, it is the need of hour to use these approaches to accelerate the okra breeding programme at faster pace. The genomics and bioinformatics should also be well integrated into the programme for effective application of markers to okra breeding. A comparative genomics approach of other crop should also be applied for breeding programmes of those crops where the genome information is not available. Development of cost-effective genotyping technologies should always be the integral part of any improvement programme. There is need to use SSR and SNP based genotyping technologies as well as advanced technologies such as next generation sequencing.

Resilient resistance to begomoviruses like Yellow vein mosaic virus (YVMV) poses a serious challenge to both breeders and pathologists as these viruses are highly diverse, and constantly generate new forms via recombination. Biotechnological tools to generate resistant cultivar against Yellow vein mosaic disease (YVMD) are limited due to the lack of informative polymorphic markers, genetic maps and genome sequence information. Therefore, use of novel molecular and genomic tools will help in the accomplishing resistance against YVMV in okra. Identification of markers linked to the YVMV resistance gene/s and its pyramiding for combining multiple disease resistance genes in various backgrounds will help in okra crop improvement. In addition, genomic tools will help in elucidating the metabolic pathway governing disease resistance.
