**3.3. Sequence-based molecular marker discovery and genetic mapping**

Sequence-based molecular markers have been used in many comparative and functional genomics studies because of their preferable features like genome-wide distribution, chromo‐ some-specific location, co-dominant inheritance, and reproducibility. The high-throughput NGS technologies produce a huge amount of data, which is highly suitable for the identifica‐ tion of a large number of sequence variations in genome or transcriptome. For SNP identifi‐ cation, various SNP calling programs such as SOAPsnp [63], MAQ [64], Atlas-SNP2 [65], SAMtools [66], and GATK [67, 68] have been used commonly [69].

In tomato, Sim et al. [70] developed the first large-scale SNP genotyping array using 8784 SNPs based on NGS-derived transcriptome sequences of six different genotypes [71]. They con‐ structed three high-density linkage maps using interspecific F2 populations (with various accessions of *S. lycopersicum* and *S. pennellii*). The physical positions of about 7666 SNPs were identified relative to the draft tomato genome sequence and found that the genetic and the physical distances were persistent. Such maps help to provide details of genetic order and recombination, also to improve gene assemblies and to dissect the complex traits. In another study, the genome-wide SNP genotyping was carried out with 7617 SNPs in 40 tomato lines and identified 6474 polymorphic SNPs [72]. Further, the effect of SNPs on protein function was studied, which revealed that the function of about 200 genes was altered by the substitu‐ tions phenomenon.

In eggplant, Barchi et al. [73] mapped QTLs associated with anthocyanin pigmentation using inter- and intraspecific linkage maps. They used a combination of the restriction site-associated DNA (RAD) strategy with high throughput sequencing (Illumina) to generate SNPs. A total of 415 of the 431 markers were assembled into twelve major and one minor linkage group, covering 1390 cM distance.

Very recently, in pepper, Devran et al. [74] developed molecular markers tightly linked to *potyvirus resistance* 4 (Pvr4) by sequencing the parental lines and progenies using Illumina Hi-Seq2500 in combination with bulked segregant analysis (BSA) approach. By comparative analysis, they identified the syntenic regions between resistant and susceptible progenies, and more than 5000 single-nucleotide variants (SNVs) were identified that were converted into CAPS markers and used to map *Pvr4* locus using F2 mapping populations. In a separate study, intron-targeting (IT) markers were developed from the NGS (5500xl SOLiD)-derived tran‐ scripts in tetraploid potato cv. White lady [75]. These markers were tested on various potato genotypes and in other *Solanum* species. A detailed list of reports of NGS-based molecular marker is given in Table 1.



more than 5000 single-nucleotide variants (SNVs) were identified that were converted into CAPS markers and used to map *Pvr4* locus using F2 mapping populations. In a separate study, intron-targeting (IT) markers were developed from the NGS (5500xl SOLiD)-derived tran‐ scripts in tetraploid potato cv. White lady [75]. These markers were tested on various potato genotypes and in other *Solanum* species. A detailed list of reports of NGS-based molecular

**SSRs**

Second assembly 10,398 22,000 Illumina

Mandarin (*C. annuum*) – 1025 454 GS-FLX [44]

BA3 (*C. annuum*) – 154,519 InDels Illumina [76]

– 1536 SNPs were

– 62,576 non

SNPs

selected for genotyping of which 1293 successfully genotyped and 1248 found polymorphic

redundant putative

TF68 (*Capsicum annuum*) 751 1536 SNPs

Blackcluster (*C. annuum*) – 1059

BA07 (*C. annuum*) – 149,755 InDels

**Number of SNPs/**

853 11,849 454 GS-FLX

2,489 4,236 Illumina [41]

4,072 9,150 Illumina [43]

– 5,000 SNV Illumina

**NGS platform**

454 GS-FLX [39]

and Illumina

HiSeq 2500

Illumina [30]

Resequencing with ABI SOLiD and Genotyping by Illumina GoldenGate Assay

[74]

[77]

**Reference**

[40]

**InDels**

101 InDels

marker is given in Table 1.

**Capsicum** 1 Transcriptome profiling

2 Transcriptome profiling

3 Transcriptome profiling

4 Transcriptome profiling

5 Transcriptome profiling

6 Whole genome resequencing

7 Genome sequencing with BSA

**Tomato** 1 Whole genome resequencing

2 Whole

transcriptome sequencing

**S. No. Type of study Population/species Number of**

258 Next Generation Sequencing - Advances, Applications and Challenges

*annuum*)

*frutescens*)

Yolo Wonder and Criollo de Morelos 334 (both *C.*

Bukang (*C. annuum*) First assembly

Xiaomila (*Capsicum*

SR231 and Criollo de Morelos334 (*C. annuum* L.)

Ailsa Craig, Furikoma,

Chuukanbonhon Nou 11, Ponderosa and Regina (All are inbred lines of *Solanum*

M82, Tomato

*lycopersicum*)

8 accessions of (*S. lycopersicum*) and 1 of (*Solanum pimpinellifolium*) Note: SNP—single-nucleotide polymorphism, SNV—single-nucleotide variant, SSR—simple sequence repeat, InDels insertion/deletion.

**Table 1.** List of transcriptome and whole genome sequencing using NGS technologies for development of genomic resources in Solanaceae crop plants

#### **3.4. Epigenomics during the age of next-generation sequencing technologies**

Molecular breeding has a crucial role in the improvement of crops. Although conventional breeding program brought a substantial increment of food production, however, with rapid population growth worldwide, crop improvement should be accelerated so that climate resilient, biotic stress-resistant, high-nutritional, and high-productivity cultivars could be developed. The advent of NGS made it possible to study phenotypic variations caused by genetic and epigenetic modification to facilitates crop improvement. The term epigenotype was first introduced by Conrad H. Waddington to demonstrate the sum of interrelated developmental pathways that enable one genome to give rise to multiple epigenomes and consequently to multiple cell types that make up the whole organism. Nowadays, the term epigenetics is commonly referred to all kinds of heritable changes that are not caused by changes in the alteration of DNA sequences but are triggered by chemical modifications on the DNA (cytosine methylation) or on histone modifications (e.g., acetylation, methylation) bringing about modulation of chromatin structure and function [83]. In recent years, small RNAs have been emerged as key players in controlling epigenetic changes throughout the plant genome.
