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DOI https://doi.org/10.18551/rjoas.2021-03.20
GENETIC BASIS OF RICE AROMA GENE AND ITS APPLICATION IN RICE GENETICS AND BREEDING: A REVIEW
Bigyan K.C.*, Pandit R., Regmi R., Bhusal B., Neupane P., Bhattarai K., Maharjan B., Acharya S.
Department of Plant Breeding, Postgraduate Program, Institute of Agriculture and Animal Science (IAAS), Tribhuvan University, Kirtipur, Nepal
Poudel Mukti Ram
Institute of Agriculture and Animal Science (IAAS), Rupandehi, Nepal
*E-mail: [email protected]
ABSTRACT
One of the most desired and highly valuable characters is aroma or fragrance, whose preference and demand is increasing day by day. Aromatic rice is considered proudly as an asset in many countries. More than 200 types of volatile substances are found in rice among which 2-acetyl-1-pyrrole (2-AP) is the most important compound in fragrance rice. By the help of map-based cloning, Badh2, the principal gene contributing 2-AP was mapped on chromosome 8. Non-functional BADH2 protein is resulted if a mutation occurs in badh2 gene. Hence, the accumulation of principal aromatic compound, 2-AP occurs in aromatic rice. This paper provides the insights into the genetic basis, gene function and application of aroma gene functional markers in the cultivation, production and breeding of aromatic rice along with its genetic improvement.
KEY WORDS
Rice aroma, genetic breeding, fragrance gene, aromatic rice, molecular mapping.
More than 3 billion population of the world depends upon the rice for 25% of energy so rice is considered one of the vital cereal crops in the world [1]. For Southeast Asians countries, it is one of the staple food crops as it supplies about 35% of their energy requirements [2, 3]. Aromatic character of rice is considered as important rice grain quality. The price of aromatic rice is increasing along with its demand. Aromatic rice are found in different parts of the world among which Basmati rice of India and Pakistan and the Jasmine type rice of Thailand are the aromatic cultivars famous in world market (Table 1).
Table 1 - Some aromatic rice found in different parts of the world [4, 5, 6]
Country Name of the aromatic rice
Afghanistan Lawangin, Barah
Bangladesh Kala Namak, Badshahbhog, Chinigura, Kalijira, Radhuni Pagal, Kataribhog, Sakkokhora
Cambodia Somali
China Xiang Kang 3, Zhao Xing 17, Xiang Nuo 4
India Pusa Basmati 1, Basmati 370, Pusa Basmati 1121, Basmati 370, CSR30, Kala Namak, Badshahbhog, Kalajeera, Dubraj, Sonachur, Ambemohar, Tilak chandan
Indonesia Rojolele, Mentek Wangi, Sukanandi
Iran Hassani, Dom Siah, Hassan Sarai, Gerdeh, Mehr, Anbar-boo, Mirza, Mosa Tarom, Salari, Sadri
Iraq Anbarboo
Japan Iwaga, Miyakaori, Kouikuka 37, Sari Queen
Myanmar Boka Hmwe, Balugyun, Nama Tha Lay, Pawsan Hmwe
Nepal Brahmphool, Rato, Jetho Budho, Jhinuwa, Achhame masino, Anadhi, Basmati, Basmati andi, Ekle, Jaran Dhan, Kalo basmati, Jogini, Kanak jira, Kasturi, Krishnabhog, Lalka Basmati, Mahabhog, Rajbhog, Sali dhan, Shyamjira, Tulsiprasad, Thapachini, etc.
Philippines Milagrosa, Azucena
Thailand Hawn Mali, Nahng Nuan, RD6, RD 15, Som Hong
Vietnam Nang Huong Ran, Di Huong, Nep Hoa Vang, Nep Rong, Tam On, Nep Rong, Tam Xoan
More than 200 types of volatile substances are found in rice among which 2-acetyl-1-pyrrole (2-AP) is the most important in fragrance rice [7]. Chewing and KOH method are mostly used in detecting aroma of rice. Genetic basis of fragrant rice is complicated, it is thought that fragrance is governed by single recessive gene (fgr), located on the eighth chromosome of rice genome. With the help of map based cloning, the principle gene badh2 has been found contributing to the 2AP [8]. In this paper we discuss the genetic basis of rice aroma gene, recent work made in study of aroma gene and application of functional markers for improvement of new varieties with high aromatic quality.
GENETIC BASIS OF AROMATIC RICE
Along with the increase in demand of aromatic rice, the study of its genetic basis has also made a lot of progress. Researchers in 1992 came with the result that the gene which controls fragrance was recessive gene, located on the chromosome number 8. Also, they succeeded in finding the genetic distance between molecular marker RG28 and aroma gene and it was 4.5 cM [9]. After that a lot of researchers also detected the fragrance controlling recessive gene by testing different types of molecular markers on different genetic population, near the RG28 (Table 2). Up to now, various reserachers have shown different markers highly associated with the fgr locus (Table 3). Zhang Tao et al. [10] succeeded in locating aroma gene about 252kb between 20 175 367 bp-20 386 172 bp on chromosome 8 by using aromatic rice and indica which is non-fragrant rice as experimental materials. And similar but deeper analysis helped in finding two molecular markers on the gene which encodes betaine dehydrogenase (BADH2). So, this finding revealed that the difference between non-aromatic and aromatic rice was governed by the difference in the sequence of BADH2 gene. After map-based cloning and sequencing of the fgr region, it was found that 7th exon region of Badh2 has mutation which disrupt the function of Badh2 protein leading to fragrance development in aromatic rice. Hence, it can be concluded that there is likelihood of linkage between Badh2 gene and fgr gene, which determine the fragrance in rice [11].
Table 2 - Molecular mapping of fgr gene
Number of genes Chromosomes Type of markers References
1 8 EST, SSR [12]
1 8 SSR [13]
2 - SSR [14]
1 8 SSR [15]
3 QTLs QTL on 8, 3 and 4 SSR [16]
1 8 SSR, RFLP, STS [17]
1 8 SSR [18]
1 8 SSR [19]
1 8 SSR [20]
The total length of Badh2 is 1509 bp including 15 exons and 14 introns [21]. Several mutations in Badh2 region was found while comparing different aromatic and non-aromatic rice. For instance, most of the aromatic rice has 8 bp deletion and 3 single nucleotide polymorphism sites (SNPs) in the 7th exon of Badh2 gene [11] (Figure 1). But 7 bp deletion in exon 2 of the Badh2 gene and 803 bp deletion between exon 4 and exon 5 has also been found in different aromatic rice varieties [22] and also variation sites were found in 1, 10, 13 and 14 exons. Further detail studies have found that the splice sites of the first exons and first introns of the Badh2 gene, the 5 'UTR region and, the promoter region also have insertion, deletion, or single nucleotide mutation sites [23]. Hence, allelic variation of Badh2 gene largely determine the difference in aroma of rice grain. On whole genome analysis of the Barth Marty's aroma gene, 2 quantitative trait loci (QTL) were found on the 3rd and 4th chromosome which control aromatic characteristics of rice in addition to the Badh2 gene [16]. The QTL located on the 4th chromosome might be related to the Badh1 gene which is homologous to Badh2 gene. Mutation in the Badh2 gene is also linked to the aromatic quality of rice but its effect is very low as compared to Badh2 gene [16]. Hence, further study is needed here.
RJOAS, 3(111), March 2021 Table 3 - Molecular markers associated with fgr locus in different populations of rice
Type of markers Linked markers Crosses Population type References
RFLP RG28 B8462T3-710/Aromatic Lemont F3 [9]
RFLP, RAPD RG28, RG1, Y5 I R64/Azucena doubled haploid (DH) line [24]
SSR RM42, RM223 a) IR64/Azucena b) Zhai-Ye-Qing 8/Jing Xi 17 c) Milyang 23/Gihobyeo d) BS 125/acc.WL02 a) DH1 line b) DH2 line c) F11 d)interspecific backcross [25]
SSR RM223, RM342A, RM515 Y-1281/Kalizira F7 [26]
Functional marker FMbadh2-E2A, FMbadh2-E2B, FMbadh2-E7 Xiangjing02-5855/Xiangxuenuo F2 [22]
SSR Aro7, RM23120, RM3459 a) Chuanxiang-29B/R2 b) Chuanxiang-29B/Lemont 2 2 FF a) b) [18]
SSR ARSSR-3 a) Taroari Basmati/B 95-1 b) Basmati 386/BPT5204 c) Vasumati/B 95-1 d) Basmati 386/BPT 5204 a) BC1F1 b) BC1F1 c) BC1F1 d) F2 [27]
SSR RM33, RM85 Milyang 23/Gihobyeo F11 [28]
SSR SCU-SSR1 Kyeema/Doongara F2 [29]
SSR SCU015RM Kyeema/Doongara F2 [30]
SSR, EST 10L03 FW, CP04133 KDML105/CT9993 F9 [12]
CAPS, STS L02 - L06 a) Wuxiangxian/Zhongai91A b) Wuxiangjing/Nanjing11 c) Suyunuo/Zhongxian3037 d) Suyunuo/Guichao2 a) BC1, F2 b) BC1, F2 c) BC1, F2, F3 d) BC1, F2 [13]
Functional co-dominant marker BADEX7-5 Basmati 386/Improved Samba Mahsuri F2 [31]
SSR, STS RM223, RG28FL-RB -- -- [17]
ASA ESP, IFAP, INSP, EAP a) Sang tarom/Neda b) Sang tarom/Nemat c) Tarom deylamani /Neda d) Tarom deylamani /Nemat 2 2 2 2 FFFF a) b) c) d) [32]
^^ Introns O Exons (2$) Promoter
Exon 7
Figure 1 - The fgr gene structure (Badh2) displaying badh2-E7 as a non-functional allele, with three SNPs and an 8 bp deletion in the 7th exon, and badh2-E2 as a second non-functional allele, including a 7 bp deletion in the 2nd exon, in aromatic rice cultivars. The non-fragrant rice cultivars with functional Badh2 alleles have 15 exons and 14 introns with an ATG start codon in the first exon and an ATT stop codon in the 15th exon [11, 22]
Several previous studies showed that the aroma gene has been transferred from earliest wild rice varieties by a long period of domestication and artificial selection [33] and hence aroma gene might have produced a range of variations during this process which resulted in the different conclusions and results among researchers. Due to this reason there is a different views regarding number of recessive gene controlling aromatic character. Most of the researchers believe that only single recessive gene control the fragrance inheritance [33, 34], while other believe it is controlled by multi recessive genes. Hence, it can be concluded that the genetic basis of rice aroma gene is complex due to different fragrant types such as jasmine, popcorn, hickory and violet and due to the environmental factors interacting with aroma genes.
AROMA GENES AND ITS APPLICATION IN RICE GENETICS AND BREEDING
Since most of the rice breeding techniques are still traditional breeding so it is laborious and time intensive to cultivate new varieties of aromatic rice using conventional breeding methods. As aroma gene is controlled by recessive gene so introduction of the Badh2 mutant gene into the existing elite varieties by hybridization and backcrossing should be done. Then, the individuals of offspring population are screened and hence, it is hard to identify the process at the seedling stage. However, Chinese rice breeders have successfully done researches on the high-yield, high-quality, multi-resistance fragrant rice varieties, and screened a series of aromatic rice restorer lines, sterile lines, and new hybrid combinations [35-37]. Molecular marker assisted selection has been widely used in accelerating the process of screening and breeding of new varieties of aromatic rice. Molecular markers such as RG28, SCU015RM, and RSP04 could differentiate between non-aromatic and aromatic rice [38]. A series of specific primers were designed by further analysing function of Badh2 gene which can screen the fragrant gene and helps in the breeding of restoration lines of aromatic lines [39]. Also, transgenic technology is used in the cultivation of new aromatic rice varieties. Transforming of non-aromatic into aromatic rice varieties can be done by RNAi mediated Badh2 gene silencing but large number of transgenic progeny plants needs to be screened because this method often does not completely inhibit the expression of Badh2 gene [40, 41].
Genome editing technology could make non-aromatic rice produce fragrance by insertion, replacement or deletion of Badh2 gene which causes premature termination codon or encoded amino acid change or even non coding corresponding Badh2 protein in rice [11, 22, 42]. Hence, it is conceivable that any mutation in Badh2 ceases its function and promote the production of 2-AP leading to the development of new aroma gene. Thus, TALEN technology which can knock out the Badh2 gene could be beneficial in transforming non-aromatic rice into aromatic rice and also can be used to create a genetically homozygous mutant of aromatic rice plant [21]. Therefore, TALEN technology greatly speeds up the breeding process of the new varieties along with other genome editing technology ZFN and CRISPR technology. These technologies are superior compare to traditional breeding as they can accurately edit the target gene and there is no need for hybridization and backcross process, thereby saving a lot of time and convenience.
CONCLUSION
Aromatic rice is high in demand and its global market value is increasing tremendously as it is highly appealing to human beings. Hence, aroma or fragrance is supposed to be a prominent trait for several breeding programs. The genetic basis of rice aroma gene is complex due to different fragrant types such as jasmine, popcorn, hickory and violet and due to the environmental factors interacting with aroma genes. Similarly, 2AP is the principle fragrant compound present in aromatic rice, several volatile components, possibly different from 2AP-associated fragrance, provides every variety its own unique aroma. However, not much is known about the association of these volatiles with aroma. Hence, further promising approach such as genome editing technology would further help in genetic improvement and breeding of aromatic rice.
RJOAS, 3(111), March 2021 CONFLICTS OF INTEREST
The authors declare that they have no conflict of interest regarding the publication of this paper.
REFERENCES
1. Tian Z, Qian Q, Liu Q, Yan M, Liu X, Yan C et al. Allelic diversities in rice starch biosynthesis lead to a diverse array of rice eating and cooking qualities. Proceedings of the National Academy of Sciences. 2009; 106(51):21760-21765.
2. Peng B. et al. OsAAP6 functions as an important regulator of grain protein content and nutritional quality in rice. Nature Communications. 2014; 5(1).
3. Li S, Gao J, Li J, Wang Y. Research Progress on the regulation of rice tillering by single legged gold lactone. Journal of plant.2015; 50.
4. Upadhyay M, Joshi B. Plant genetic resources in SAARC countries: Their conservation and management: Nepal chapter. SAARC Agricultural Information Centre (SAIC), BARC complex, Dhaka-1215, Bangladesh.
5. Khush G. In Aromatic Rices: Oxford & IBH Publishing Company, New Delhi; 2000.
6. Sarkarung S, Somrith B, Chitrakorn S. In Aromatic Rices: Oxford & IBH Publishing Company, New Delhi; 2000
7. Mathure S. et al. Comparative quantitative analysis of headspace volatiles and their association with BADH2 marker in non-basmati scented, basmati and non-scented rice (Oryza sativa L.) cultivars of India. Food Chemistry. 2014; 142:383-391.
8. Siddiq E, Vemireddy L, Nagaraju J. Basmati Rices: Genetics, Breeding and Trade. Agricultural Research. 2012; 1(1):25-36.
9. Ahn S, Bollich C, Tanksley S. RFLP tagging of a gene for aroma in rice. Theoretical and Applied Genetics. 1992; 84-84(7-8):825-828.
10. Zhang T, Zhang H , Jiang K, Xu P, Wang X, Wu X., Zheng, J. Fine mapping of rice flavor genes. Molecular Plant Breeding. 2008; 6; 1038-1044.
11. Bradbury L, Fitzgerald T, Henry R, Jin Q, Waters D. The gene for fragrance in rice. Plant BiotechnolJ. 2021; 3: 363-370.
12. Wanchana S. et al. A rapid construction of a physical contig across a 4.5 cM region for rice grain aroma facilitates marker enrichment for positional cloning. Sci. Asia. 2005; 31: 299-306.
13. Chen S, Wu J, Yang Y, Shi W, Xu M. The fgr gene responsible for rice fragrance was restricted within 69kb. Plant Science. 2006; 171(4):505-514.
14. Hien N, Yoshihashi T, Sarhadi W, Thanh V, Oikawa Y, Hirata Y. Evaluation of Aroma in Rice (Oryza sativa L.) using KOH Method, Molecular Markers and Measurement of 2-Acetyl-1-Pyrroline Concentration. Nettai nogyo Jpn. J. Tropical Agric. 2006; 50:190-198.
15. Li J, Wang F, Liu W, Jin S, Liu Y. Genetic analysis and mapping by SSR marker for fragrance gene in rice Yuefeng B. Mol. Plant Breed. 2006; 4: 54-58.
16. Amarawathi Y, Singh R, Singh A, Singh V, Mohapatra T, Sharma T et al. Mapping of quantitative trait loci for basmati quality traits in rice (Oryza sativa L.). Molecular Breeding. 2007; 21(1):49-65.
17. Lang N, Buu B. Initial Marker-Assisted Selection in Rice Breeding at Cuu Long Delta Rice Research Institute. Omonrice. 2010; 17: 8-21.
18. Sun S, Gao F, Lu X, Wu X, Wang X, Ren G et al. Genetic analysis and gene fine mapping of aroma in rice (Oryza sativa L. Cyperales, Poaceae). Genetics and Molecular Biology. 2008; 31(2):532-538.
19. Bradbury L. Identification of the gene responsible for fragrance in rice and characterisation of the enzyme transcribed from this gene and its homologs. PhD thesis, Southern Cross University, Lismore, Australia. 2009.
20. Yi M, Nwe K, Vanavichit A, Chai-arree W, Toojinda T. Marker assisted backcross breeding to improve cooking quality traits in Myanmar rice cultivar Manawthukha. Field Crops Research. 2009; 113(2):178-186.
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
Shan Q, Zhang Y, Chen K, Zhang K, Gao C. Creation of fragrant rice by targeted knockout of theOsBADH2gene using TALEN technology. Plant Biotechnology Journal. 2015; 13(6):791-800.
Shi W, Yang Y, Chen S, Xu M. Discovery of a new fragrance allele and the development of functional markers for the breeding of fragrant rice varieties. Molecular Breeding. 2008; 22(2):185-192.
Ootsuka K, Takahashi I, Tanaka K, Itani T, Tabuchi H, Yoshihashi T et al. Genetic polymorphisms in Japanese fragrant landraces and novel fragrant allele domesticated in northern Japan. Breeding Science. 2014; 64(2):115-124.
Lorieux M. et al. Aroma in rice: genetic analysis of a quantitative trait. TAG Theoretical and Applied Genetics. 1996; 93(7):1145-1151.
Chen X, Temnykh S, Xu Y, Cho Y, McCouch S. Development of a microsatellite framework map providing genome-wide coverage in rice (Oryza sativa L.). Theoretical and Applied Genetics. 1997; 95(4):553-567.
Kibria K, Islam M, Begum S. Screening of aromatic rice lines by phenotypic and molecular markers. Bangladesh Journal of Botany. 1970; 37(2):141-147. Mahdav M, Pandey M, Kumar P, Sundaram R, Prasad G, Sudarshan L, Rani S. Identification and mapping of tightly linked SSR marker for aroma trait for use in marker assisted selection in rice. Rice Genet. Newsl. 2009; 25: 38-39.
Cho Y, McCouch S, Kuiper M, Kang M, Pot J, Groenen J et al. Integrated map of AFLP, SSLP and RFLP markers using a recombinant inbred population of rice (Oryza sativa L.). Theoretical and Applied Genetics. 1998; 97(3):370-380.
Garland S, Lewin L, Blakeney A, Reinke R, Henry R. PCR-based molecular markers for the fragrance gene in rice (Oryza sativa. L.). Theoretical and Applied Genetics. 2000; 101(3):364-371.
Cordeiro G, Christopher M, Henry J, Reinke R. Identification of microsatellite markers for fragrance in rice by analysis of the rice genome sequence. Mol. Breed. 2002; 9: 245-250. Sakthivel K. et al. Development of a simple functional marker for fragrance in rice and its validation in Indian Basmati and non-Basmati fragrant rice varieties. Molecular Breeding. 2009; 24(2):185-190.
Kiani G. Marker aided selection for aroma in F2 populations of rice. African Journal of Biotechnology. 2011; 10(71).
He Q, Yu J, Kim T, Cho Y, Lee Y, Park Y. Resequencing Reveals Different Domestication Rate for BADH1 and BADH2 in Rice (Oryza sativa). PLOS ONE. 2015; 10(8):e0134801.
Shao G, Tang S, Chen M, Wei X, He J, Luo J et al. Haplotype variation at Badh2, the gene determining fragrance in rice. Genomics. 2013; 101(2):157-162. Huang T. et al. Breeding and utilization of restorer line Daolixiang 15 of indicarice. Fujian Journal of Agricultural Sciences. 2006; 21: 83-88.
Liu G, Lu X, Ren G, Gao F, Li Z, Ren M, Tang J. Breeding and cultivation techniques of a new hybrid rice combination Chuanxiangyou 425. Chinese Rice. 2006; 2:42-43. Jiang Q, Lin G, Zhao D, Li Y, He B, Wang F. Characteristics and utilization of aromatic high quality sterile line Yixiang 1A. Chinese Rice. 2008; 2: 35-37.
Jin Q. et al. A single nucleotide polymorphism (SNP) marker linked to the fragrance gene in rice (Oryza sativa L.). Plant Sci. 2003; 165: 359-364.
Yan Y, Zhu G, Zhang L, Wan C, Cao L, Zhao Z, Wu S. Development and application of rice flavor gene molecular markers. Northwestern Journal of Botany. 2005; 35:269-274. Chen M, Wei X, Shao G, Tang S, Luo J, Hu P. Fragrance of the rice grain achieved via artificial microRNA-induced down-regulation ofOsBADH2. Plant Breeding. 2012; 131(5):584-590.
Peng B, Sun Y,Chen B, Sun R, Kong D, Pang R, Song S et al. Research progress of fragrance gene and its application in rice breeding. Chin Bull Bot.2017; 52: 797-807. Kovach M, Calingacion M, Fitzgerald M, McCouch S. The origin and evolution of fragrance in rice (Oryza sativa L.). Proceedings of the National Academy of Sciences. 2009; 106(34): 14444-14449.
