Doubled haploid (DH) is a condition formed when haploid cells undergo chromosome sdoubling. Artificial production of doubled haploids is important in crop improvement. Double haploid (DH) breeding not only helps in accelerating conventional plant breeding programmes and make early release of cultivars with superior and desirable traits possible but it has greater utility in research aspects of plant breeding, genetics and genetic engineering. DHs are important constituent of germplasm. The production of haploids and doubled haploids(DHs) through gametic tissues allows a single-step development of complete homozygous lines from heterozygous parents. DHs shorten the time required to produce homozygous plants in comparison with the conventional breeding, which requires several generations of selfing (Mishra et al., 2021).

By the early 1980s, androgenesis had been reported in 171 species, many of which were important crop plants. Gynogenesis was reported in 15 species, in which androgenesis was not successful. The value of these haploids was that they could be used to detect mutations and for recovery of unique recombinants, because there is no masking of recessive alleles. As well, the production of double haploids allowed for hybrid production and their integration into breeding programs. Haploids are generated by in vitro procedures based on the culture of immature male and female gametophytes and by in vivo procedures based on inter and intraspecific hybridization causing uniparental chromosome elimination. Once haploid plants become available, their genome must be doubled to produce a fertile DH line (Ren et al., 2017).

The study was conducted to relate flower bud size with microspore developmental stages and the culture media had concentration of plant growth regulator effects on the in vitro androgenesis. Flower buds were randomly collected and visually divided into three stage based on both petal and sepal size. Then, the anthers were cultured on MS basal medium with different concentration of NAA (0.1 - 0.7 mg/L), kinetin (1.0 - 3.0 mg/L), and BA (0.5 - 1.5 mg/L). The results showed that anthers with light violet, in colour and 2.5 mm long, consisted of anthers with 80% haploid and 20% dihaploid microspores in the first mitosis were selected. In induction culture media, it was observed that MS medium with 2.0 mg/L Kinetin and 0.5 mg/L NAA gave embryos at higher frequencies. MS medium with 1 mg/L BA is the best medium for embryo germination and inducting shoots. (Phuong., 2021)

The results of the investigation conducted by Olszewska and Nowaczyk in 2021 proved the gametophytic origin of diploids derived from two-embryonic seeds of pepper C. annuum L. hybrid. During the germination of seeds harvested from the red fruited hybrid (C.

annuum L. ATZ × C. annuum L. ‘Sono’) F1 , additional embryos have been found. Four of diploid twins F2 generation were different in their phenotype within the pair and/or from the F1 mother plant. Plants of three pairs: 1A–1B, 2A–2B, 4A–4B were significantly different with regard to the average fruit weight, length and seeds number. Yellow colour of ripe fruit was characteristic for 2A, 3A and 3B plants. In RAPD molecular analysis, twenty three primers were used and six of them enabled polymorphic products to be obtained in reactions. The results of the analysis confirmed phenotypic differentiation of the twins and their parental forms. The phenotypic and molecular analyses proved that spontaneous diploids from a two-embryonic seeds are ready for the production of genetically stable, sexual progeny. (Olszewska and Nowaczyk., 2021)

Genetic mapping using DHs had an impact in locating gene control traits for yield, quality, agronomy, abiotic and biotic stress. DHs now feature in cultivar production in a number of crops, and breeding time is considerably reduced. Hence, worldwide use of DH technology as an accelerated approach to crop improvement has become routine by many breeding companies and laboratories leading to the development of further new varieties.

REFERENCES:

Mishra, A. K., Saini, R. and Tiwari, K. N., 2021, Double haploid production and its applications in crop improvement. Agril. Biotech.: Latest Research and Trends : 75-101. 

Olszewska, D. and Nowaczyk, P., 2021, Double haploids from two-embryonic seeds of pepper (Capsicum annuum L.) F. Acta Sci. Pol. Hortorum Cultus, 20(3): 45–52.

 Phuong, N. T. D. 2021, Direct embryogenesis from anther culture of hot chilli Capsicum annuum L. Ho chi minh city open university journal of science-engineering and technology, 11(1): 3-10.

Ren, J., Wu, P., Trampe, B., Tian, X., Lübberstedt, T. and Chen, S., 2017, Novel technologies in doubled haploid line development. Pla. Bio., 15(11): 1361-1370.