Advances and Milestones of Radish Breeding: An Update

Abstract

Radish is a member of the Cruciferae family. The important traits for radish breeding include high yield, early maturity, late bolting, pungency, cold-hardiness, drought resistance, heat tolerance and soil adaptability. For successful radish production, one needs to the understand nature and behaviour of the flower and very important to identify the S haplotypes of parental lines to produce F1 hybrids based on self-incompatibility to get rid of laborious hand emasculation in radish. Therefore, further breeding programmes depend on inter-specific and intra-specific hybridization, which is vital in genomic studies and crop improvement by introducing desirable agronomic characters. It is essential to acquire detailed genetic information on chromosomes and inheritance. Genomics is now at the core of radish breeding to study the underlying differences in genotypes. Moreover, researchers have produced transgenic radishes with various agronomic characteristics over the last decade.

1. Introduction

Radish is an annual herbaceous vegetable known as Raphanus sativus [1], and it is a diploid containing two sets of 18 chromosomes (2n = 18) [2]. Radish belongs to the Cruciferae family and is eaten fresh as grated radish, a garnish and a salad [3]. Radish is regularly served in eastern Asian cuisine; radish has also featured in food worldwide [4]. There is a focus on developing high-quality radish varieties ideal for tropical and subtropical temperatures [5]. Breeding work has been performed on numerous agronomical traits including tolerance to pathogens and consumption adaptability. Traits for radish breeding include high yield, early maturity, late bolting, pungency, cold-hardiness, drought resistance, heat tolerance and soil adaptability [3]. There is a positive correlation between the radish’s consistency and its amount of sugar, pungency, elaboration of the cell, water content and pore extent [6, 7]. Although, the main endeavour has been to modify the radish cultivation to various growing seasons [8]. It is essential to acquire detailed genetic information on chromosomes and information on inheritance for multiple genes responsible for agronomical, biochemical traits and resistance to biotic and abiotic stresses for carrying out a successful radish breeding [9, 10, 11]. Marketing assessment and consumer preference are primarily associated with physical attractiveness such as length, shape, size and skin colour [12]. A primary colour changes into white and different pink, red, purple, yellow and green.

The anthocyanin pelargonidin is the colour-causing pigment in red colour radish varieties, and they have a mild flavour (not as pungent) and are around 40 cm in length [13]. Quality-related traits are remarkably heritable. They are often strongly influenced by cultivation methods. The swollen tap roots of radishes may be oval, tapered or cylindrical [14]. Moreover, mechanical harvesting often includes cylindrical root cultivars [15]. Rich in antler velvet, radish roots produce useful phytochemicals. They have cancer-preventive properties and a significant contributor to the taste and flavour of Brassica vegetables [16]. In addition, radishes provide complex carbohydrates, dietary fibre often organic nutrients and minerals to humans [17].

Omics approaches using Next-Generation Sequencing (NGS) methods provide a large amount of genomic data that enable the identification of novel genes and sequences. In addition, genome-wide study results reveal the genetic causes of diverse characteristics [18, 19]. Furthermore, less study has been published that discusses the historical milestones and technological advancements in radish breeding. As a result, we have gathered information on different aspects of radish breeding and its numerous accomplishments in this section. We believe that this work will prove to be a valuable resource for vegetable breeders in the future.

2. Breeding goals

Radish is high in its nutrition content, health benefits conferred by its chemical compounds and a significant contribution to the human being [20]. Root length is an important trait of radish for consumers, and preference always goes to radish on length, diameter and colour; visual indicators such as the colour of the label and where it is presented are crucial to buyers’ selection phase. Too many experiments have concentrated on gene mapping to classify significant colour-gene associations in various vegetables for this cause [21]. Productive radish root colour is vital to radish crop productivity. The discovery and detection of considerable plant genes involved in radish colouring would aid in advancing colour genetics [22]. The range of potential skin inheritance trends has been investigated extensively in radish. It was reported that 609 chemical compounds are present within 23 categories of the most studied varieties of Radish, such as red (30%), white (13%) and black (6%) [23]. This study also found that nutrients, antioxidants and phytochemicals are mainly identified in roots, sprouts and leaves, which could be considered an important part of a healthy diet [23]. In addition, researchers have focused on red radish with red flesh because it contains large amounts of a natural red pigment widely used in foods, wine and cosmetics. The uniformity of various colours, sizes and yields are the factors becoming a high-priority goal in radish breeding [24]. Radish has a wide variety of colours that affect its appearance and its nutritional quality [25]. However, the detection, identification and quantification of flavonoids in multicolour radish are rarely explored. At the same time, it was also identified that red and purple radishes contained similar anthocyanin compounds responsible for colour pigmentation, including red cyanidin, callistephin and pelargonin [26]. Purple ZJL contains cyanidin o-syringic acid and cyanin, whereas callistephin and pelargonin contain more amount in dark red TXH. The metabolites in coloured radishes that differed from SZB genes are broadly involved in the plant secondary metabolites biosynthetic pathway, such as flavonoid, flavone, isoflavonoid and phenylpropanoid biosynthesis. This approach would be useful for cultivating important and valuable new radish varieties [26, 27]. These results explain anthocyanin synthesis in radish and provide potential genetic clues for improving anthocyanins in radish roots [28].

Fusarium wilt (FW) is a soil-borne vascular wilt disease caused by fungal pathogen Fusarium oxysporum f. sp. Raphanin, causes severe yield losses in radish production [29]. The most effective method to control the FW is using resistant varieties in crop improvement. Fusarium resistance is highly studied among ‘Motohashi-’ or ‘Kuroba-mino’ lines of the Minowase variety, and Tosai’ is the strongest line among Nerami varieties [30]. A pathogen could damage harassment of yield, and in root colour, these varieties would not be preferred for the consumer. Bioactive compounds in R. sativus (radish) are being studied to treat several diseases. Therefore, radish has attracted scientific attention due to its nutritional and phytochemical composition, which reduces the risk of developing many cancers and cardiovascular diseases. Further, the important goals are provided in Figure 1.

Figure 1.

Breeding goals of radish (Raphanus sativus L.).

Moreover, salinisation is considered as one of the significant soil pollutions in the environment affecting plant growth and soil fertility globally [31]. This scenario alarms an urgent need to enrich the soil or to identify stress-tolerant plants. It is reported that antioxidant enzymes (HOD-Hydrogen Peroxide; SOD-Superoxide; LOD-Lipid Peroxidation; CAT-Catalase) play a major role in reducing the effects of salts in plants by monitoring the oxidative stress in them [32]. To study the salt tolerance of Japanese wild radishes called ‘Hamadaikon’ (R. sativus f. raphanistroides Makino) and its characteristics such as seed germination, plant height, root length and fresh weight were examined under the salinity condition. It was found that higher germination and growth in NaCl were shown at 25°C than those at 20°C [33]. Hence, wild radishes could be considered for salt tolerance breeding. Moreover, halopriming is a seed priming technique in which the seeds were soaked in various salt solutions to enhance germination and seedling emergence uniformly under adverse conditions. The effect of halopriming on germination, initial growth and development of radish under salt stress conditions was studied. It was found that the best outcome was achieved by priming with CaCl₂ for germination characteristics and vigour and with KCl for initial development [34].

3. Botany of radish

Radish (R. sativus L.) is an entomophilous flower classified as an allogamous plant [35]. Regular flowering appears from three florets on the tip of each branch of the panicle, and each flower is effective in producing a pod up to 1–3 inches long and consists of 1–6 seeds. The radish flowers open in the morning with fresh corolla and remain until the next day [36]. Kremer also reported that pollen receptivity of the flower limits up to few hours a day. Its flowers are 1.5–2 cm in width, whitish to pinkish and purplish colour with purple veins and have four erected sepals and clawed petals, six stamens and 3–4 cm long style [37, 38]. Siliqua or seedpod, a type of seed capsule of radish, is 1.5 cm wide and 3–7 cm long, consisting of 6–12 seeds/pod with a long conical seedless beak [13]. The inflorescence of radish is a typical elongated, erected, an oblong raceme of Cruciferae. The main objective of the investigation was the cross-pollination of radish by [39] found that the ‘Icicle’ and ‘Scarlet Globe’ cvs were self-incompatible pollinated with the help of honeybees [40]. The studies indicated that the seed yield is greatly influenced by the number of honeybees striking the radish flowers. Radchenko [41] also reported that honeybees were the main pollinators of radish flowers, approximately 77–99% in total, increasing the crop yield by 22% and enhancing the seed quality. Therefore, radish is considered almost entirely insect-pollinated. During fruit maturation, seeds’ colour is somewhat yellow and turns reddish-brown with age. The mature radish leaves are alternate, arranged in a rosette pattern and have a lyrate shape set apart pinnately with an enhanced terminal lobe and minor lateral lobes. A longer root form, including oriental radishes, daikon or mooli and winter radishes, grows up to 60 cm (24 in) long with foliage about 60 cm (24 in) high with a spread of 45 cm (18 in) [42].

4. S haplotype

Radish is a self-incompatible crop exhibiting the high heterosis, the production of F₁ hybrids based on self-incompatibility is desired to eliminate laborious hand emasculation in radish [43]. The main aim of a plant breeder is to identify breeding lines of S haplotypes. The plant breeder can avoid cross-influencing of the parental lines [44]. The S haplotype of each parental line needs to show the compatible reaction between parental lines. Therefore, for producing F₁ hybrid breeding, it is very important to identify the S haplotypes of parental lines [45]. The abundance of S haplotype determines a specific S haplotype by using traditional methods, including test cross method, pollination, isoelectric focusing, immunoblot analysis and the pollen tube fluorescence analysis [46, 47]. The S alleles are highly variable in S haplotype. Moreover, Nikura and Matsuura identified 37 alleles in radish [48].

Several S haplotypes in Raphinus sativus were identified based on polymorphism in SLG, SRK and SCR/SP11 sequence and S haplotypes are numbered as S-1, S-2, S-3, etc. [48]. Although radish belongs to a genus different from Brassica, nucleotide sequences of SP11, SRK and SLG alleles of radish and Brassica are intermingled in phylogenetic trees of SP11, SRK and SLG, respectively, indicating that diversification of these alleles predates speciation of these genera [44]. SP11, SRK and SLG alleles of some S haplotypes in radish are highly similar to those of some S haplotypes in Brassica, and one S haplotype in radish has been revealed to have the same recognition specificity as that of one S haplotype in Brassica rapa [44]. Comparison of nucleotide sequences of SP11 and SRK alleles and recognition specificities between similar S haplotypes of radish and Brassica may provide valuable information for understanding the molecular structures of SP11 and SRK proteins. However, researchers’ numbering of S haplotypes in radish varies, and nucleotide sequence information on S haplotypes is thus confusing [43].

Besides, analysis of SLG and SRK is utilised to identify S haplotype in Raphanus and Brassica by using methods of polymerase chain reaction–restriction fragment length polymorphism (PCR-RFLP) [49, 50]. However, the own limitation of PCR-RFLP, first, is challenging to design a universal primer that can amplify SLG and SRK alleles; second, the presence of multiple genes homologous to the SLG or SRK genes in Brassicaceae plants aggravates PCR amplification of specific SLG or SRK alleles [51, 52, 53]. Additional advanced radish cultivars (cultivars with improved yield and higher quality) were also produced by the Ogura CMS method to assist in radish hybrid [54]. This variety shows that bulk selection, mixed mass pedigree selection or bud pollination will take 8–12 years to produce a new variety, new varieties must be produced from other genetic means [55].

5. Inter-specific hybridization

Inter-specific and intra-specific hybridization has a vital role in the genomic study and crop improvement by introducing desirable agronomic characteristics and specific traits such as disease, insects and stress resistance from wild species to cultivated ones [56, 57]. The study indicated that the average podding rate of the cross between radish and turnip (67.03%) was much higher than that of the reciprocal cross between a turnip and radish (55.04%) [57], and it was also reported that the average seed-setting rate and hybrid acquisition rate of the radish and turnip based on cross pattern (e.g. 2.25 and 0% respectively), however, seed production of the F₁ hybrids and their F₂ progeny was up to 0.4 and 2%, respectively, as compared with wild radish [58]. Therefore, the study indicated a low hybridization affinity between radish and Chinese kale, but incompatibility still prevailed [57].

Similarly, the radish-wild mustard inter-specific hybrid was studied. It was found that production was higher with radish pollen competition, i.e. 42 and three interspecific hybrid seeds per 1000 seeds were observed [59]. Another study indicated that the modified flower culture method is the best method for hybridization between radish and (transgenic) oilseed rape (Raphanobrassica hybrids) without labour-intensive production in vitro ovule or embryo rescue techniques. This is a potential approach for breeding programmes by introducing useful radish genes, e.g. nematode resistance genes, into oilseed rape [60]. Moreover, clubroot is a common disease of cabbages, cauliflower, radishes, turnips and other plants of the family Brassicaceae caused by Plasmodiophora brassicae [61]. Radish is a close relative of the brassica family, and it was found that a synthesised allotetraploid Brassicoraphanus (RRCC, 2n = 36) between R. sativus cv. HQ-04 (2n = 18, RR) and Brassica oleracea var. alboglabra (L.H Bailey) (2n = 18, CC) proved resistant to multiple clubroot disease pathogen P. brassicae causing club root disease [62]. However, the spontaneous hybridization event between Brassica napus (oilseed rape) and Raphanus raphanistrum (wild radish) was screened. It was found that hybrids with wild radish as the seed parent contribute to herbicide resistance belonging to rape. Another study indicated that wild radish in an oilseed rape field produced as many as three interspecific hybrids per 100 plants and was the first ever such report of such a spontaneous event [58].

6. Molecular markers to QTLian breeding

Several economically important traits in radish are being identified. These traits are yield, insect resistance and disease resistance. Yield is a complex trait governed by polygenic characters. Identifying these traits using conventional breeding/traditional breeding is problematic because these traits depend on phenotypic expression and have environmental and genotypic interaction. The new tool molecular breeding overcomes these problems, identification of quantitative trait is being utilised with the help of DNA markers [7] and linkage mapping [63]. There are several DNA markers being used in breeding programmes, such as restriction fragment length polymorphism (RFLPs), random amplified polymorphism DNA (RAPD), simple sequence repeats (SSRs), single-nucleotide polymorphism (SNPs) [63, 64, 65, 66, 67]. Molecular markers such as RAPD have been used to establish the origin of hortensis var. sativus and var. niger, which formed from distinct progenitors and came from different sources [68]. Several Asian varieties show more incredible skin and flesh colour, size, length and weight of roots, var. hortensis’ genetic variability is also not a surprise.

A genome-wide association study (GWAS) analysis identified 44 single-nucleotide polymorphisms (SNPs) and 20 putative candidate genes significantly associated with FW resistance. A total of four QTLs were identified from the F₂ population derived from an FW resistant line and a susceptible line, one of which was co-located with the SNPs on chromosome 7. These markers are emerging tools for molecular breeding programmes and marker-assisted selection to develop FW-resistant varieties of R. sativus [69]. Moreover, for the identification of the disease Fusarium wilt, Yu et al. [70] constructed a genetic linkage map on the F₂ population, they observed a total of eight loci conferring FW resistance that were distributed on 4LGs, namely 2, 3, 6 and 7 of the Raphanus genome. Synteny analysis using the linked markers QTL showed homology to A. thaliana chromosome 3, which contains disease-resistance gene clusters, suggesting the conservation of resistance genes between them. The list of significant QTLs is identified in the radish, and their location is provided in Table 1.

Locus ID	Locus Name	Trait	Cross	Population	Marker Name	Max LOD	References
t3726.T000001	qFW1	Fusarium wilt resistance	835 x B2	F2	nu_mBRPGM1376	3.72	70
t3726.T000002	qFW1	Fusarium wilt resistance	835 x B2	F2	ACMP0606	4.34	70
t3726.T000003	qFW1	Fusarium wilt resistance	835 x B2	F2	nu_mBRPGM1208	9.42	70
t3726.T000004	qFW1	Fusarium wilt resistance	835 x B2	F2	ACMP0357	10.08	70
t3726.T000005	qFW1	Fusarium wilt resistance	835 x B2	F2	nia032a	3.31	70
t3726.T000006	qFW2	Fusarium wilt resistance	835 x B2	F2	nu_mBRPGM1376	3.29	70
t3726.T000007	qFW3	Fusarium wilt resistance	835 x B2	F2	ACMP0590	5.62	70
t3726.T000008	qFW4	Fusarium wilt resistance	835 x B2	F2	nu_mBRPGM0432	3.76	70
t3726.T000009	qFW5	Fusarium wilt resistance	835 x B2	F2	nu_mBRPGM0432	4.84	70
t3726.T000010	qFW6	Fusarium wilt resistance	835 x B2	F2	nu_mBRPGM0432	4.23	70
t3726.T000011	qFW7	Fusarium wilt resistance	835 x B2	F2	ACMP0606	7.94	70
t3726.T000012	qFW8	Fusarium wilt resistance	835 x B2	F2	ACMP0606	8.84	70
t3726.T000013		Root shape (length/width)	Huang-he hong wan x Utsugi-gensuke	F2	BRMS-303-2	2.42	65
t3726.T000014		Thickening	Huang-he hong wan x Utsugi-gensuke	F2	ACA/CTT5	3.03	65
t3726.T000015		Thickening	Huang-he hong wan x Utsugi-gensuke	F2	AAG/CTA12	4.51	65
t3726.T000016		Root shape (length/width)	Huang-he hong wan x Utsugi-gensuke	F2	ACA/CTG5	2.88	65
t3726.T000017		Red pigmentation	Huang-he hong wan x Utsugi-gensuke	F2	SLG	9.58	65
t3726.T000017		Red pigmentation	Huang-he hong wan x Utsugi-gensuke	F2	SLG	9.58	65
t3726.T000018		CR (clubroot resistance), Crs1 (clubroot resistance locus of Rap	Koga-benimaru x Utsugi-gensuke	F2	BN142	12.64	71
t3726.T000019	qRCd1	RCd (mg kg^−1), root Cd concentration	Nau-Dysx x Nau-Yh	F2	Ni2E05_525	5.24	63
t3726.T000019	qRCd1	RCd (mg kg^−1), root Cd concentration	Nau-Dysx x Nau-Yh	F2	Ni2E05_525	5.24	63
t3726.T000020	qRCd4	RCd (mg kg^−1), root Cd concentration	Nau-Dysx x Nau-Yh	F2	BRMS129_510	4.36	63
t3726.T000020	qRCd4	RCd (mg kg^−1), root Cd concentration	Nau-Dysx x Nau-Yh	F2	BRMS129_510	4.36	63
t3726.T000021	qRCd6	RCd (mg kg^−1), root Cd concentration	Nau-Dysx x Nau-Yh	F2	EM6fc3_331	5.28	63
t3726.T000021	qRCd6	RCd (mg kg^−1), root Cd concentration	Nau-Dysx x Nau-Yh	F2	EM6fc3_331	5.28	63
t3726.T000022	qRCd9	RCd (mg kg^−1), root Cd concentration	Nau-Dysx x Nau-Yh	F2	EM5me6_286	23.64	63
t3726.T000022	qRCd9	RCd (mg kg^−1), root Cd concentration	Nau-Dysx x Nau-Yh	F2	EM5me6_286	23.64	63
t3726.T000023	qRDW5	RDW (g), root dry weight	Nau-Dysx x Nau-Yh	F2	BRMS058_550	7.28	63
t3726.T000024	qRDW6	RDW (g), root dry weight	Nau-Dysx x Nau-Yh	F2	NAUrp705_644	3.96	63
t3726.T000025	qRDW9	RDW (g), root dry weight	Nau-Dysx x Nau-Yh	F2	EM3me6_291	3.58	63
t3726.T000026	qRL1	RL (cm), root length	Nau-Dysx x Nau-Yh	F2	Na10F06_545	3.38	63
t3726.T000027	qRL3.1	RL (cm), root length	Nau-Dysx x Nau-Yh	F2	RamRM24-568_618	3.26	63
t3726.T000028	qRL3.2	RL (cm), root length	Nau-Dysx x Nau-Yh	F2	PM2fc8_314	3.64	63
t3726.T000029	qRL5	RL (cm), root length	Nau-Dysx x Nau-Yh	F2	NAUrp782_643	4.89	63
t3726.T000030	qRL7	RL (cm), root length	Nau-Dysx x Nau-Yh	F2	EM4odd48_365	4.06	63
t3726.T000031	qSCd1	SCd (mg kg^−1), shoot Cd concentration	Nau-Dysx x Nau-Yh	F2	NAUrp362_706	4.37	63
t3726.T000031	qSCd1	SCd (mg kg^−1), shoot Cd concentration	Nau-Dysx x Nau-Yh	F2	NAUrp362_706	4.37	63
t3726.T000032	qSCd3	SCd (mg kg^−1), shoot Cd concentration	Nau-Dysx x Nau-Yh	F2	EM16ga18_383	7.64	63
t3726.T000032	qSCd3	SCd (mg kg^−1), shoot Cd concentration	Nau-Dysx x Nau-Yh	F2	EM16ga18_383	7.64	63
t3726.T000033	qSDW2	SDW (g), shoot dry weight	Nau-Dysx x Nau-Yh	F2	NAUJKC19_565	4.74	63
t3726.T000034	qSDW6	SDW (g), shoot dry weight	Nau-Dysx x Nau-Yh	F2	Na10F08_665	3.78	63
t3726.T000035	qSDW9	SDW (g), shoot dry weight	Nau-Dysx x Nau-Yh	F2	Ol14E06_720	4.62	63
t3726.T000036	qSH2	SH (cm), shoot height	Nau-Dysx x Nau-Yh	F2	EM16ga18_384	4.25	63
t3726.T000037	qSH5	SH (cm), shoot height	Nau-Dysx x Nau-Yh	F2	RGA12F12R_300	3.64	63
t3726.T000038	qTDW1.1	TDW (g), total dry weight	Nau-Dysx x Nau-Yh	F2	RamM2-706_616	3.54	63
t3726.T000039	qTDW1.2	TDW (g), total dry weight	Nau-Dysx x Nau-Yh	F2	PM2em11_367	5.6	63
t3726.T000040	qTDW5	TDW (g), total dry weight	Nau-Dysx x Nau-Yh	F2	PM17em10_327	4.43	63
t3726.T000041	qTDW6	TDW (g), total dry weight	Nau-Dysx x Nau-Yh	F2	NAUrp586_752	3.14	63
t3726.T000042	qTDW7	TDW (g), total dry weight	Nau-Dysx x Nau-Yh	F2	PM18odd44_362	4.84	63
t3726.T000043	qTDW9	TDW (g), total dry weight	Nau-Dysx x Nau-Yh	F2	PM36em8_423	6.2	63
t3726.T000044	Hs1_Rph	BCN-resistance, resistance against the beet cyst nematode (H. sc	Pegletta x Siletta Nova	F2	E41M59–297	22.6	66
t3726.T000045		PS, Plant shape	rat-tail radish x Haru-S	RIL	RsHH019	3.8	67
t3726.T000046		MW, Main root weight (g)	rat-tail radish x Haru-S	RIL	REL-13	6.9	67
t3726.T000047		Pubescence	rat-tail radish x Haru-S	RIL	RM_1	10.1	67
t3726.T000048		MW, Main root weight (g)	rat-tail radish x Haru-S	RIL	RES-1	11.7	67
t3726.T000049		WW, Whole plant weight (g)	rat-tail radish x Haru-S	RIL	AtSTS-1015	3.8	67
t3726.T000050		Pubescence	rat-tail radish x Haru-S	RIL	RsSR104	13.6	67
t3726.T000051	GSL-QTL-1	4MTB-GSL contents, 4-methylthio-3-butenyl glucosinolate contents	TBS x AZ26H	F2	RS2CL6432s	5.87	72
t3726.T000052	GSL-QTL-1	4MTB-GSL contents, 4-methylthio-3-butenyl glucosinolate contents	TBS x AZ26H	F2	S2CL4585s	3.62	72
t3726.T000053	GSL-QTL-1	4MTB-GSL contents, 4-methylthio-3-butenyl glucosinolate contents	TBS x AZ26H	F2	RS2CL3356s	3.85	72
t3726.T000054	GSL-QTL-1	4MTB-GSL contents, 4-methylthio-3-butenyl glucosinolate contents	TBS x AZ26H	F2	RS2CL4290s	7.36	72
t3726.T000055	GSL-QTL-2	4MTB-GSL contents, 4-methylthio-3-butenyl glucosinolate contents	TBS x AZ26H	F2	RS2CL6432s	19.1	72
t3726.T000056	GSL-QTL-3	4MTB-GSL contents, 4-methylthio-3-butenyl glucosinolate contents	TBS x AZ26H	F2	RS2CL6594s	5.62	72
t3726.T000057	GSL-QTL-4	4MTB-GSL contents, 4-methylthio-3-butenyl glucosinolate contents	TBS x AZ26H	F2	RS2CL6594s	1.54	72
t3726.T000058	GSL-QTL-5	4MTB-GSL contents, 4-methylthio-3-butenyl glucosinolate contents	TBS x AZ26H	F2	S2CL4585s	5.19	72

Table 1.

List of QTLs identified for important traits in radish.

Moreover, identification of root shape and red pigmentation is performed, and it was observed that three quantitative trait loci for root shape, namely LG3, LG8 and LG9, two QTLs for root diameter, namely LG4 and LG8 and one for red pigmentation are identified with the help of using AFLP, SSR and SLG-CAPS [65]. Kamei et al. [71] constructed a genetic linkage map using AFLP and SSR markers and concluded that CR is governed by the single gene or closely linked gene loci, namely Crs1, Crs2 and Crr3. A genetic map was constructed using an F₂ population by using markers SRAP, RAPD, SSR, ISSR, RAMP and RGA markers, and they found that a novel QTL qRCD9 is responsible for controlling root CD [63]. Resistance against cyst nematode (Heterodera schachtii) was identified using RAPD, dpRAPD, AFLP and SSR markers [66]. To identify quantitative traits in radish for morphological characters, namely ovule number per silique, seed number per silique, plant shape, pubescence, whole plant weight (g), upper part weight (g), whole root weight (g), main root weight (g) using recombinant inbred lines, they identified 8 and 10 quantitative traits in 2008 and 2009 respectively [67]. In the identified QTL regions, nine SNP markers were newly produced. Nucleotide sequences and expression of these genes suggested their possible function in 4MTB-GSL biosynthesis in radish roots. [72]. Whereas recently, it was discovered that the R2R3-MYB transcription factor responsible for anthocyanin pigment 2 (PAP2) production is located on chromosome 2. The amino acid sequence encoded by the RsPAP2 gene was entirely distinguishable from other previously published RsMYB genes responsible for the red skin colour of radish [73].

7. Genomics

Genomics is now at the core of crop improvement, and the radish crop has been exploited to study the underlying differences in genotypes. The rapid development of genomic data boosted the discoveries regarding the genetic basis of plant traits, such as increased yield, flowering or disease resistance [74]. Various studies of the radish have investigated their genomes’ arrangement and the reorganisation of chromosomes during polyploidy events [75], of which draft genomic sequences have been assembled. Another study reported an Asian radish cultivar, WK10039 which was sequenced entirely by combining 454, Illumina and PacBio sequencing systems and bacterial artificial chromosome clones obtained through end sequencing was fully sequenced by using the end sequencing method and sequencing equipment from the ABI firm [76]; over the last decade, a variety of genomic studies on the cultivated radish have been performed [77]. Moreover, a chromosome-scale genome assembly (rs1.0) of WK10039, an Asian radish cultivar, was constructed compared with assemblies documented previously [78]. It revealed more details than those previously recorded (having greater coverage of the genome, a greater number of contigs and chromosome anchoring) [79]. However, Radish Base is a genomic and genetic database containing radish mitochondrial genome sequences [80]. This database presently includes the mitochondrial genomes of two newly sequenced radish species, one from the normal cytoplasm and the other from the male-sterile cytoplasm of Ogura [81]. The previous study published the bioinformatics analysis in radish and identified 20 COL transcription factors in the radish genome among 54,357 coding genes [82]. Every COL gene in the ‘Aokubi daikon’ cultivar matched the COL gene in the ‘kazusa’ cultivar. A total of 20 radish COL genes were also searched in the cultivar ‘WK10039’ [82]. Besides, in the radish genome, 35 unique RsOFPs and five RsOFP-likes (with no/partial OVATE domain) were identified by BLASTP, and analysis of exon-intron organisation revealed that most genes were intron-less containing maximum coding sequences in the genome [82].

Based on 17-mer analysis, the estimated size of the genome came out to be 530 Mb. A 387.73 Mb was assembled into 44,820 high-quality scaffolds using SOAP denovo [83, 84] and SSPACE [85]. The assembly in this study showed excellent results with fosmid clones (98.86% covered). The assembly showed a much higher quality than the draft genome of Raphanus raphanistrum (254 Mb contigs) [86] and two assemblies (116.0 and 179.8 Mb) of R. sativus ‘Aokubi’ [87] which was released previously. After de novo assembly of the ‘Okute-Sakurajima’ genome, an estimated haploid genome size of 498.5 Mb was found. The de novo assembly showed a substantially heterozygous genome [88]. Subsequent long-read sequencing produced 36.0 Gb data (60.7 coverage of the estimated genome size) in 2.3 million reads with an N50 length of 29.1 kb. After two rounds of data polishing, the long-read assembly consisting of 504.5 Mb primary contigs (including 1437 sequences with an N50 length of 1.2 Mb) and 263.5 Mb alternative contigs consists of the other haplotypes with different alleles, also known as haploid sequences (including 2373 sequences with an N50 length of 154.6 kb) [88].

A study performed on the radish genome after polyploidy has shown fundamental information about the radish genome production and evolution, which provides valuable insights into radish genetics and breeding. The detailed data and genomic methods obtained through these investigations support a greater understanding of the radish triplicated genome composition. Additionally, these methods help radish breeding by promoting marker-assisted collection, comparative genomic studies and the transmission of knowledge from the reference data to other radish accessions [89]. Consequently, a portal that is home to considerable quantities of genomic information and various links to specific genome analysis methods is precious in radish research and breeding.

8. Genetic engineering

Genetic engineering has pivotal role in agriculture by improving the characteristics in the crops and satisfying the need of poor nourished countries. The developments in gene technology and metabolic engineering systems accelerated the production of valuable germplasms [90]. Progress is being achieved in plant methods by improving the traits; researchers have successfully produced transgenic radishes with various agronomic characteristics [91, 92, 93]. Gene transmission is done with the help of pathogen, known as agrobacterium, which is extensively used for plant hairy root lines, which appear to yield better than other forms of root systems [94]. Herbaceous hairy roots have advantageous due to their longevity, pace of growth and capacity to assist plants in growing from the root up [95]. The hairy roots are produced in nutrient solution with the help of increasing agrobacterium that contains unusual properties, including biochemically and bio-transforming different metabolites. It is best to use Agrobacterium to produce secondary metabolites since they help to enhance growth regulators [96]. Working on the hairy roots, new sources of natural compounds [97]. In addition, chromosomal disruption or amplification may affect the fertility of cultivated plants. Antibiotics, herbicides, metabolic analogues and non-toxic agents all facilitate transformed cells for survival. Kanamycin and hygromycin B hamper radish regeneration [98].

Recent advancements in plant biotechnology indicate that radish could be genetically modified via a process called ‘floral-dipping’. This technique involves co-suppression of the photoperiodic gene GIGANTEA in radish and contributes to the plant’s ability to delay bolting and blooming. It can be used to boost a crop’s medicinal value [98]. The prospects for improving transformation efficiency and selecting new traits for generating late-flowering radish are published [68]. In 2001, it was demonstrated that plants derived from plants dipped into an Agrobacterium suspension containing both the beta-glucuronidase (gusA) gene and the herbicide resistance gene (bar) between the flanking T-DNA border sequences could be used to generate transgenic radish (R. sativus L. longipinnatus Bailey) [91]. In the end, Southern blotting results revealed that both the gusA and bar genes integrated into the genome of transformed plants and segregated as dominant Mendelian traits [91]. A study revealed that The RHA2b gene from radish encodes a transcription factor involved in abscisic acid (ABA) signal transduction and is responsible for seed dormancy and pre-harvest sprouting [99]. The study performed the experimentation in which The RsRHA2b gene was cloned and transferred into Zhengmai 9023 via Agrobacterium-mediated stem apex transformation [99]. The agrobacterium-mediated transformation became a more appropriate method for genetic transformation [100]. Using adventitious shoot growth on hypocotyl explants for Agrobacterium-mediated radish genetic transformation was investigated using transgenic radish (Raphanus sativa L., cv. Jin Ju Dae Pyong) grown on Murashige and Skoog medium [101]. Besides, northern blot results revealed that the GUS gene transcript was detected in a few regenerated plants, confirming genetic transformation. In addition, the techniques available for introducing pharmaceutical proteins into radish for on-site delivery of edible proteins into it are discussed by Curtis in his study [98]. The concerns of releasing transgenic radish to the field in pollen-mediated gene transfer have also been explored. Risks that might exist and the introduction of transgenic radish to the field are sometimes brought up in discussions about transgenic crops [91, 102, 103].

9. Conclusion and future directions

For successful production of radish yield, inter- and intra-specific hybridizations are vital to genetic research and crop improvement because they enable the introduction of desirable agronomic traits into the population. The production of yields, early maturity and late bolting, pungency, cold-hardiness, drought resistance, heat tolerance and soil adaptability are just a few of the essential radish breeding traits. The radish genome contains self-incompatibility alleles, allowing for the generation of F₁ hybrids without the labour-intensive and hand emasculation required in radish. When generating F₁ combinations, it is critical to determine the S haplotypes of the parental lines to avoid hand emasculation. Collecting complete genetic data on chromosomes and information on inheritance is critical. To better understand and forecast resistance, yield characteristics and fruit quality, researchers must understand the regulatory factors synchronising at various developmental stages for each attribute discussed. It remains necessary to develop a robust and long-lasting strategy for plant disease resistance, which is currently under consideration. This is because diseases are capable of evading resistance by generating novel bacterial strains.

Speed breeding is one such strategy; as genome sequencing costs continue to decline, RAD-sequencing and DNA microarrays will become more common, enabling faster genome mapping and tagging of new quantitative trait loci. These quantitative trait loci (QTLs) may incorporate resistance into high-yielding radish genotypes and combine them with significant resistance genes to increase the number of resistant radish genotypes. Additionally, GWAS (genome-wide association studies) can map characteristics to specific candidate genes on a genome-wide scale to improve crop production and quality in radish. The discovery of significant genetic and metabolic diversity paves the way to develop controlled harvest variations in agriculture and genetic enhancement via breeding.

Funding

This research has received no external funding.

Conflict of interest

Authors declare no conflict of interest.