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Author(s): P. B. Ingle1, S. S. Rokade*2, R.V.Raut3

Email(s): 1, 2rokadesim@gmail.com, 3

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    1S.S.S.K.R Innani Mahavidyalaya, Karanja (Lad), Dist. Washim (M.S.) India
    2Department of Botany Late Pundalikrao Gawali Arts and Science Mahavidyalaya, Shirpur (Jain) Dist. Washim (M.S.) India
    3Nirmal Seeds Pvt.Ltd. Pachora Dist. Jalgaon (M.S.) India
    *Corresponding Author Email- rokadesim@gmail.com

Published In:   Volume - 5,      Issue - 1,     Year - 2023


Cite this article:
P. B. Ingle, S. S. Rokade, R.V.Raut (2023) Review on double haploid in rice plant (Oryza sativa L.). NewBioWorld A Journal of Alumni Association of Biotechnology,5(1):1-4.

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 NewBioWorld A Journal of Alumni Association of Biotechnology (2023) 5(1):1-4 

REVIEW ARTICLE

Review on double haploid in rice plant (Oryza sativa L.)

P. B. Ingle1, S. S. Rokade2*, R.V.Raut3

 

1S.S.S.K.R Innani Mahavidyalaya, Karanja (Lad), Dist. Washim (M.S.) India.

2Department of Botany Late Pundalikrao Gawali Arts and Science Mahavidyalaya, Shirpur (Jain) Dist. Washim (M.S.) India.

3Nirmal Seeds Pvt.Ltd. Pachora Dist. Jalgaon (M.S.) India.

rokadesim@gmail.com

*Corresponding Author Email- rokadesim@gmail.com

ARTICLE INFORMATION

 

ABSTRACT

Article history:

Received

14 April 2023

Received in revised form

27 May 2023

Accepted

29 May 2023

Keywords:

Homozygous; Androgenesis;

Doubled haploid;

Rice;

Plant Breeding

 

Rice (Oryza sativa L.) is an important and indigenous crop in India. Rice is a plant belonging to the family of grasses, Gramineae (Poaceae). It is a major food crop of the world and forms the staple diet of about half of the world's population. Rice is repeatedly mentioned in ancient Indian texts, including the Yajur Veda. The area under cultivation of rice in India is 45416 thousand hectare, Total production of Rice during 2020-21 is estimated at record 122.27 million tonnes. Two major Asian cultivated varieties of rice have been historically recognized as japonica and indica subspecies. Japonica rice is sticky and indica rice is non sticky. Obtaining high quality homozygous pure lines by classical breeding methods is very time consuming and labour intensive. Due to uncontrolled pollination, the results are hard to obtain through this method. In-vitro haploid production offers an excellent solution for development of high quality homozygous pure lines in lesser time. Indica rice is mainly cultivated in tropical and subtropical environments at lower latitudes or altitudes, whereas japonica rice is grown mainly in more temperate environments at higher latitudes or altitudes. Anther culture is easy in japonica rice, but its application in Indica rice is limited as it is difficult to breed through anther culture. Indica cultivars of rice showed low anther culturability (1.2% callus induction) whereas Japonica cultivars had 20-fold higher (28.1%) anther culturability.

 


Introduction

DOI: 10.52228/NBW-JAAB.2023-5-1-1

Rice (Oryza sativa L.) is an important and indigenous crop in India. Rice is a plant belonging to the family of grasses, Gramineae (Poaceae). It is a major food crop of the world and forms the staple diet of about half of the world's population. Archaeological evidence suggests that this grain was the basis of India’s ancient civilizations. The rice is supposed to have been cultivated in the valley of the Ganges River as far back as 6500 BC. India is the second-largest producer of rice in the world, and it also is among the largest consumers of this grain. Around over 50% of India’s people depend on rice for sustenance. Rice is repeatedly mentioned in ancient Indian texts, including the Yajur Veda. It is closely associated with fertility and health across India’s many cultural traditions. (Asia Society 2023). The area under cultivation of rice in India is 45416 thousand hectare, Total production of Rice during 2020-21 is estimated at record 122.27 million tonnes. It is higher by 9.83 million tonnes than the last five years' average production of 112.44 million tonnes (Annual Report 2021-22). Rice is a highly important cereal in the world, with a total of 490.9 million tonnes produced in 2015-16 of which more than 80% was destined to direct human consumption (FAO Trade and Market Division 2017). Rice is supposed to have undergone single domestication event around ∼8,200–13,500 year ago, estimated on the basis on the molecular clock with the help of resequencing 630 gene fragments on chromosomes 8, 10, and 12 that is supported through Demographic modeling based on SNP data and a diffusion-based approach (Molina, J et al., 2011). Higher plants show two alternations of generations i.e. sporophytic and gametophytic generation. The sporophytic form is somatic (soma=body) having a complete set of chromosomes usually diploid (2n) and the gametophytic form is haploid (n) in genomic constitution. Haploid plants have gametophytic (n) chromosomes and doubled-haploid (DH) plants have sporophytic (2n) chromosome numbers. The haploid plants derived from a diploid are known as monoploid, while a haploid plant derived from a polyploid is a polyhaploid. Haploid plants occur spontaneously or haploid can be induced by in-vivo or in-vitro methods (Dwivedi et al., 2015). Natural sporophytic haploids in the higher plants were first spotted in Jimson weed (Datura stramonium L.) (Blakeslee et al., 1922). Guha and Masheswari made the first breakthrough in the production of pollen embryogenesis by successfully producing anther culture of Datura innoxiaat the Department of Botany in the University of Delhi, India (Guha and Masheswari 1964). The rice (Oryza sativa) genome is composed of 12 chromosomes (2n = 24) which has a total length of 430 Mb. (Kurata et al., 1994). 95% of the genome of rice is mapped and it can be studied as a model plant for the grasses. (International Rice Genome Sequencing Project, 2005). The anther culture technique was first developed by Niizeki and Oono in rice (Niizeki and Oono 1968).

Homozygous pure lines

Obtaining high quality homozygous pure lines by classical breeding methods is very time consuming and labour intensive. Due to uncontrolled pollination, the results are hard to obtain through this method. In-vitro haploid production offers an excellent solution for development of high quality homozygous pure lines in lesser time. In-vitro methods for haploid plant production shorten time up to a year by using androgenesis to obtain haploid plants through tissue culture. Microspore embryogenesis is an important and useful culture technique to obtain full homozygous lines from only the male gametes. Plant regenerated with a single chromosome set, originating from microspores of in-vitro cultivated anthers, are ideal for genetic analysis due to the variety of possible expressions of the genetic makeup. The doubling of the haploid genome results in fully homozygotic lines. It shortens the period which is important for creation of genetic diversity. It also forms a base of new varieties with higher quality for crop development.

The use of haploid technique among modern breeding techniques provides tremendous advantages for breeders, especially since it allows the production of 100% homozygous pure lines only in one generation (Kurtar et al. 2010). After a haploid plant is produced, the plant can undergo diplodizaiton (2n), where the chromosome number is doubled upon stimulation with colchicines (Mohammadi et al. 2007). This method is advantageous when compared to the alternative method and is used by plant breeders or institutions around the world. Doubled haploid (DH) technology allows for the production of pure lines, useful for plant breeding through a one-generation procedure that considerably reduces the time and resources needed to produce them. The doubled haploid technique aims to generate pure inbred lines for basic research and as commercial plant varieties. The doubled haploid technique first generates haploid plants which is followed by chromosome doubling. DH Technology is of core importance in breeding programs, since it allows the production of true breeding (pure) lines, homozygous for all traits, in a way faster and cheaper than classical breeding procedure. This reduces considerably the number of generations needed to produce a pure line, thus decreasing the cost of breeding programmes with increase in variability in germplasm.

Because traditional breeding activities cannot respond quickly to market mobility but with the integration of dihaploidization methods into the breeding cycles, breeding programs have gained significant momentum. Present efforts are an attempt to standardize development of an efficient in-vitro development of double haploid in Rice crop using anther culture.  Two major Asian cultivated verities of rice have been historically recognized as Japonica and Indica subspecies. Japonica rice is sticky and Indica rice is non sticky (Yanget al., 2014). Indica rice is mainly cultivated in tropical and subtropical environments at lower latitudes or altitudes, whereas Japonica rice is grown mainly in more temperate environments at higher latitudes or altitudes. Anther culture is easy in Japonica rice, but its application in Indica rice is limited as it is difficult to breed through anther culture. Indica cultivars of rice showed low anther culturability (1.2% callus induction) whereas Japonica cultivars had 20-fold higher (28.1%) anther culturability (Grewal et al., 2011) because of early anther necrosis, poor callus proliferation and regeneration of albino plantlets (Chen et al, 1991).

For a long time, conventional breeding methods have been used to obtain pure, 100% homozygous lines for hybrid seed production in crops of agronomic interest. However, by double haploid technology, it is possible to produce 100% homozygous plants derived from precursors of male gametophytes (androgenesis), to accelerate the production of pure lines, which implies important time and cost savings. Producing homozygous breeding lines via in-vitro haploid techniques are much more efficient in terms of time and cost as compared to other methods (Bajaj et al., 1990). Anther culture protocol in eggplant is being already reported till regeneration and acclimation of doubled haploid plants (Calabuig et al., 2020).Adopting the method of haploid identification, from undesired heterozygote plants will substantially affect the success of androgenesis in breeding programs (Ahmadi & Ebrahimzadeh, 2020).The future for double haploid breeding is promising as vigorous DH protocols are available for a growing number of crops and future applications will witness a closer integration with molecular-marker and gene-splicing technologies (Humphreys and Knox, 2015).

Ploidy analysis

Ploidy analysis is done by flow cytometry and SSR based PCR markers. Flow cytometry allowed rapid and reliable determination of ploidy level in anther culture derived plant (Martínezet al., 1994). Microspore-derived plants are analyzed by flow cytometry to check their DNA contents and by comparison with a known diploid individual, infer their ploidy level. Thus, in parallel to microspore derived plants, diploid donor plants are analyzed as control for the 2C DNA content to compare with flow cytometry. SSR (simple sequence repeat) markers are considered as the key choice for genotyping. They provide highly reproducible results as compared to the other PCR-based markers (Song et al., 2019).  Simple sequence repeats (SSRs) are abundant and are distributed throughout the rice genome. They can be suitably used for understanding the source of origin of callus i.e. originated from the microspore mother cells or embryogenesis from diploid somatic tissue. SSRs have been successfully used to identify homozygous spontaneous-doubled haploids in rice hybrids (Samantaray et al., 2021).

Significance of double haploid line

Studies on haploid and double haploid line development have been carried out at very few places in India and the work is abandoned in remaining parts of the country for commercialization to the farmer community. There is continually a prerequisite to carry intensive and extensive studies on this aspect using in-vitro laboratory conditions. It will help in the generation of variable germplasm to meet the requirement of food in the coming days of increasing population in India and worldwide.

References

Ahmadi B, & Ebrahimzadeh H (2020) In-vitro androgenesis: Spontaneous vs. artificial genome doubling and characterization of regenerants. Plant cell reports39(3):299-316.

Asia society (2023). India Rice https://asiasociety.org/korea/rice-india#:~:text=Has%20rice%20always%20been%20a,far%20back%20as%206500%20BC. Accessed on 23-05-2023.

Bajaj Y P S (1990) In-vitro production of haploids and their use in cell genetics and plant breeding. In Haploids in crop improvement (pp. 3-44). Springer, Berlin, Heidelberg.

Blakeslee A F, Belling J, Farnham ME, & Bergner, AD (1922) A haploid mutant in the jimson weed," Datura stramonium". Science55(1433): 646-647.

Calabuig-Serna A, Porcel R, Corral-Martínez P, & Seguí-Simarro JM (2020) Anther culture in eggplant (Solanum melongena L.). In Plant Embryogenesis (pp. 283-293). Humana, New York, NY.

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