SNP Molecular Markers Research Articles

Genetic Diversity Analysis of Capsicum frutescens Based on Simplified Genome Sequencing Technology

Capsicum frutescens (C. frutescens) has rich germplasm resources, but there have been no reports on its genetic diversity analysis alone using simplified genome sequencing technology (GBS). To provide a basis for the breeding of new pepper varieties and the later development of C. frutescens SNP molecular markers, this study used GBS technology to sequence 65 collected pepper germplasm resources. A total of 1,399,391 SNP sites were obtained by GBS simplified genome sequencing, and there were 1,465,897 SNP variant sites. Through population genetic structure analysis, the population structure and phylogenetic tree of 65 C. frutescens germplasms were constructed. The GBS method is also suitable for the genetic relationship analysis of C. frutescens, and it also shows that there is an obvious separation of materials from different origins, and there is also a certain degree of genetic exchange. Most of C. frutescens varieties from Fujian Province and its surrounding areas are clustered together; C. frutescens varieties from western China are also clustered together. We selected T62 and T60 with a genetic distance of 0.2796 and a spiciness level of nine as the female and male parent combinations, respectively, and bred a new high-yield C. frutescens combination, ‘Mingjiao 308’. C. frutescens varieties from the same geographical origin are usually clustered together. These germplasms may contain the ancestry of multiple varieties. This result can also provide basic data for the later construction of an SNP fingerprint database.

DArTseq-Based, High-Throughput Identification of Novel Molecular Markers for the Detection of Blackleg (Leptosphaeria Spp.) Resistance in Rapeseed.

Blackleg disease, caused by Leptosphaeria spp. fungi, is one of the most important diseases of Brassica napus, responsible for severe yield losses worldwide. Blackleg resistance is controlled by major R genes and minor quantitative trait loci (QTL). Due to the high adaptation ability of the pathogen, R-mediated resistance can be easily broken, while the resistance mediated via QTL is believed to be more durable. Thus, the identification of novel molecular markers linked to blackleg resistance for B. napus breeding programs is essential. In this study, 183 doubled haploid (DH) rapeseed lines were assessed in field conditions for resistance to Leptosphaeria spp. Subsequently, DArTseq-based Genome-Wide Association Study (GWAS) was performed to identify molecular markers linked to blackleg resistance. A total of 133,764 markers (96,121 SilicoDArT and 37,643 SNP) were obtained. Finally, nine SilicoDArT and six SNP molecular markers were associated with plant resistance to Leptosphaeria spp. at the highest significance level, p < 0.001. Importantly, eleven of these fifteen markers were found within ten genes located on chromosomes A06, A07, A08, C02, C03, C06 and C08. Given the immune-related functions of the orthologues of these genes in Arabidopsis thaliana, the identified markers hold great promise for application in rapeseed breeding programs.

Genetic diversity analysis and core germplasm bank construction in cold resistant germplasm of rubber trees (Hevea brasiliensis)

The rubber tree, Hevea brasiliensis (Willd. ex Adr. de Juss.) Muell. Arg., is the sole plant worldwide utilized for the commercial production of natural rubber. Following years of breeding, there exists a wide array of germplasm differentiation in rubber trees. The exploration of diversity and population structure within rubber tree germplasm resources, alongside the establishment of core germplasm resources, is instrumental in elucidating the genetic background and facilitating the effective utilization and management of these resources. By employing SNP molecular marker technology, 195 rubber tree resources were amplified, their genetic diversity analyzed, and a fingerprint map was subsequently constructed. Through this process, the cold-resistant core germplasm of rubber trees was identified. The results revealed that the PIC, He, and pi values ranged from 0.0905 to 0.3750, 0.095 to 0.5000, and 0.0953 to 0.5013, respectively. Both group structure analysis and cluster analysis delineated the accessions into two groups, signifying a simple group structure. A core germplasm bank was established with a sampling ratio of 10%, comprising 21 accessions divided into two populations. Population G1 consists of 20 accessions, while population G2 comprises 1 accession. The research findings have led to the creation of a molecular database that is anticipated to contribute to the management and subsequent breeding applications of rubber tree accessions.

Zinc finger knuckle genes are associated with tolerance to drought and dehydration in chickpea (Cicer arietinum L.).

Chickpea (Cicer arietinum L.) is a very important food legume and needs improved drought tolerance for higher seed production in dry environments. The aim of this study was to determine diversity and genetic polymorphism in zinc finger knuckle genes with CCHC domains and their functional analysis for practical improvement of chickpea breeding. Two CaZF-CCHC genes, Ca04468 and Ca07571, were identified as potentially important candidates associated with plant responses to drought and dehydration. To study these genes, various methods were used including Sanger sequencing, DArT (Diversity array technology) and molecular markers for plant genotyping, gene expression analysis using RT-qPCR, and associations with seed-related traits in chickpea plants grown in field trials. These genes were studied for genetic polymorphism among a set of chickpea accessions, and one SNP was selected for further study from four identified SNPs between the promoter regions of each of the two genes. Molecular markers were developed for the SNP and verified using the ASQ and CAPS methods. Genotyping of parents and selected breeding lines from two hybrid populations, and SNP positions on chromosomes with haplotype identification, were confirmed using DArT microarray analysis. Differential expression profiles were identified in the parents and the hybrid populations under gradual drought and rapid dehydration. The SNP-based genotypes were differentially associated with seed weight per plant but not with 100 seed weight. The two developed and verified SNP molecular markers for both genes, Ca04468 and Ca07571, respectively, could be used for marker-assisted selection in novel chickpea cultivars with improved tolerance to drought and dehydration.

Development of seven SNP molecular markers based on the key genes related to red colour of fruit skin in Japanese apricot (Prunus mume)

Development of seven SNP molecular markers based on the key genes related to red colour of fruit skin in Japanese apricot (Prunus mume)

Genetic diversity and genome-wide association analysis of pine wood nematode populations in different regions of China.

Pine wilt disease (Bursaphelenchus xylophilus) was recently detected in Liaoning Province, which was previously considered an unfavourable area for B. xylophilus due to its low temperatures. This study aims to compare the reproductivity and genetic variations of B. xylophilus isolates from Liaoning Province and other parts of China to explore their phenotypic and genomic differences. The samples from Liaoning, Anhui, Hubei, Henan, Zhejiang and Jiangsu were isolated and purified to obtain the strains. The reproductivity of the strains was determined at 15 °C. The genetic structure was analyzed by using SNP molecular markers, and the whole genome association analysis was carried out by integrating SNP information and feculence traits. A reproductivity experiment showed that Liaoning isolates have higher reproductive ability at 15 °C. Subsequent SNP profiling and population differentiation analysis revealed obvious genetic differentiation of Liaoning isolates from other isolates. A genome-wide association study showed that SNPs closely related to low-temperature tolerance were mainly located in GPCR, Acyl-CoA, and Cpn10, which are responsible for adaptation to environmental factors, such as temperature change. Pine wood nematodes likely adapted to the climate in Liaoning and maintained a certain reproductive capacity at low temperature via variants of adaptation-related genes. This study provides a theoretical basis for elucidating the prevalence and diffusion status of B. xylophilus in China.

Transcriptome, Ectopic Expression and Genetic Population Analysis Identify Candidate Genes for Fiber Quality Improvement in Cotton.

Comparative transcriptome analysis of fiber tissues between Gossypium barbadense and Gossypium hirsutum could reveal the molecular mechanisms underlying high-quality fiber formation and identify candidate genes for fiber quality improvement. In this study, 759 genes were found to be strongly upregulated at the elongation stage in G. barbadense, which showed four distinct expression patterns (I-IV). Among them, the 346 genes of group IV stood out in terms of the potential to promote fiber elongation, in which we finally identified 42 elongation-related candidate genes by comparative transcriptome analysis between G. barbadense and G. hirsutum. Subsequently, we overexpressed GbAAR3 and GbTWS1, two of the 42 candidate genes, in Arabidopsis plants and validated their roles in promoting cell elongation. At the secondary cell wall (SCW) biosynthesis stage, 2275 genes were upregulated and exhibited five different expression profiles (I-V) in G. barbadense. We highlighted the critical roles of the 647 genes of group IV in SCW biosynthesis and further picked out 48 SCW biosynthesis-related candidate genes by comparative transcriptome analysis. SNP molecular markers were then successfully developed to distinguish the SCW biosynthesis-related candidate genes from their G. hirsutum orthologs, and the genotyping and phenotyping of a BC3F5 population proved their potential in improving fiber strength and micronaire. Our results contribute to the better understanding of the fiber quality differences between G. barbadense and G. hirsutum and provide novel alternative genes for fiber quality improvement.

Discovery of SNP Molecular Markers and Candidate Genes Associated with Sacbrood Virus Resistance in Apis cerana cerana Larvae by Whole-Genome Resequencing Based Selective Sweep Analysis

Discovery of SNP Molecular Markers and Candidate Genes Associated with Sacbrood Virus Resistance in Apis cerana cerana Larvae by Whole-Genome Resequencing Based Selective Sweep Analysis

Development of SNP molecular markers associated with resistance to reniform nematode in soybean using KASP genotyping

Reniform nematode (Rotylenchulus reniformis, Linford and Oliveira) is a sedentary, semi-endoparasite that infects a wide range of plant hosts and is one of the top three nematode pathogens affecting soybean in the southeastern United States. Previous studies have linked resistance to reniform nematode in soybean to two quantitative trait loci on chromosomes 11 and 18. A Kompetitive Allele-Specific PCR (KASP) assay was designed using SNP markers within these two regions to distinguish reniform nematode-resistant soybean based on genotype. A collection of 44 soybean plant introductions with resistant phenotype to reniform nematode and 40 susceptible soybean lines were genotyped at the two target loci to validate the KASP assay design. Of the 44 observed resistant lines, two carried the susceptible genotype; PI 438489B at the locus on chromosome 18 and PI 495017C on chromosome 11. Of the 40 observed susceptible soybean lines, only 25 had the expected susceptible genotype at the loci on chromosome 18 and 13 on chromosome 11. Our KASP assay was 68% accurate in predicting the phenotype of 84 soybean accessions based on their genotype at the SNP marker on chromosome 18 and 83% accurate at chromosome 11. These results indicate a moderate correlation of soybean SNP markers GlyREN18_46 and GlyREN11_190 with reniform nematode resistance. Further research is required to improve the accuracy of KASP assays to predict soybean response to reniform nematode, particularly host susceptibility.

Genetic Diversity Analysis of Non-Heading Chinese Cabbage of Resistance to Clubroot Disease Based on SNP Molecular Markers

Clubroot disease is a kind of soil-borne disease that seriously infects Brassica species. In this study, we collected 121 varieties of non-heading Chinese cabbages. In order to better understand the genetic variation and to screen suitable clubroot disease-resistant parental material, we re-sequenced them to examine the population genetic structure, population genetic diversity, population differentiation index, and selective sweep based on SNPs. The mapping rate with the reference genome was high, and data quality analysis revealed that the sequencing quality was good. The annotated data indicated that intronic and intergenic areas held the majority of SNPs and indels. Four subgroups of 121 non-heading Chinese cabbages were identified using principal component analysis, phylogenetic tree, and genetic structure analysis. An examination of genetic diversity revealed that while selfing may happen in subgroups C and D, heterozygosity may exist in subgroups A and B. In subgroup B, self-fertilization is not possible. There was a moderate degree of genetic differentiation between subgroups B and C (Fst = 0.0744347). For genes in certain sweep regions, we also ran GO enrichment and KEGG enrichment analysis. Two disease resistance-related genes, BraA01g042910.3.5C and BraA06g019360.3.5C, were examined. These findings will serve as a theoretical foundation for developing novel, clubroot disease-resistant types of non-heading Chinese cabbages.

Analysis of the genetic structure and diversity of upland cotton groups in different planting areas based on SNP markers.

Genetic diversity, kinship and population genetic structure analyses of Gossypium hirsutum germplasm can provide a better understanding of the origin and evolution of G. hirsutum biodiversity. In this study, adopt 273 elite upland cotton cultivar accessions collected from around the world, and especially from China to get 1,313,331 SNP molecular markers, it were used to construct a phylogenetic tree of each sample using MEGAX, to perform population structure analysis by ADMIXTURE software and principal component analysis (PCA) by EIGENSOFT software, and to estimate relatedness using SPAGeDi. ADMIXTURE software divided the experimental cotton population into 16 subgroups, and the Gossypium hirsutum samples could be roughly clustered according to source place, but there were some overlapping characteristics among samples. The experimental cotton population was divided into six groups according to source to calculate the genetic diversity index (H), and the obtained value (0.306) was close to that for germplasm collected by others in China. Cluster 4 had a relatively high genetic diversity level (0.390). The degrees of genetic differentiation within the experimental cotton population groups were low (the population differentiation indexes ranged from 0.02368 to 0.10664). The genetic distance among cotton accessions varied from 0.000332651 to 0.562664014, with an average of 0.25240429. The results of this study may provide a basis for mining elite alleles and using them for subsequent association analysis.

Development of Two Functional Markers of &lt;i&gt;Badh2&lt;/i&gt; Gene in Guangxi Fragrant Rice

Fragrance in rice is one of the most important quality traits, which resulted from the loss of function of betaine aldehyde dehydrogenase ( Badh2 ) gene on chromosome 8. The mutation of&nbsp; Badh2 &nbsp;leads to accumulation of 2-acetyl-1-pyrroline (2-AP), which is known as the main volatile in fragrant rice. At least 18 allelic variations have been identified in&nbsp; Badh2 &nbsp;genes in rice. Marker assisted selection has proved to be an effective way of fragrant rice breeding. Traditional marker detection methods, such as Sanger sequencing or SSR molecular marker, are found to be low efficient. To develop a more dynamic method, we adopt Real-time PCR method to detect the common mutation site in Guangxi. In this study,&nbsp; Badh2 &nbsp;gene of 40 fragrant rice accessions collected from Guangxi province was sequenced. Most of the fragrant rice accessions belonged to 806 bp deletion between exon 4-5 ( badh2-E4-5.1 ) and 8 bp deletion in exon 7 ( badh2-E7 ). Two Real-time PCR SNP molecular markers were developed, and were used to verify the 40 sequenced fragrant rice accessions. 93.75% of the detection results using Real-time PCR were consistent with the results of Sanger sequencing. Further, 50 local varieties were examined by Real-time PCR. A total of 24 accessions carry&nbsp; badh-E4-5&nbsp; allele and 22 accessions detected with&nbsp; badh-E7 . The two functional SNP molecular markers common in Guangxi fragrant rice were developed and proved to be useful in rice breeding. These functional markers will improve the efficiency of fragrant rice breeding.

Genomic and transcriptomic analyses of Citrus sinensis varieties provide insights into Valencia orange fruit mastication trait formation

Valencia orange (Citrus sinensis Osbeck) (VO) is a type of late-ripening sweet orange whose ripening occurs 4 to 5 months later than that of the mid-ripening common sweet orange (CO). Notably, the mastication trait of VO fruit is inferior to that of CO fruit. To date, how inferior pulp mastication trait forms in VO has not been determined. In this study, 13 VO varieties and 12 CO varieties were subjected to whole-genome resequencing. A total of 2.98 million SNPs were identified from 25 varieties, and a SNP molecular marker was developed to distinguish VO and CO. Moreover, 144 and 141 genes identified by selective sweep analysis were selected during VO and CO evolution, respectively. Based on gene functional enrichment analysis, most of the selected VO genes were related to the stress response and lignin biosynthesis. Simultaneously, we comparatively analyzed the transcriptome profiles of peel and pulp tissues among three VO varieties and three CO varieties, and the results demonstrated differences in lignin biosynthesis between VO and CO fruits. Furthermore, coexpression network analysis was performed to identify hub genes of lignin-related and variety-specific networks, which included CsERF74, CsNAC25, CsHSFB3, CsSPL4/13, etc. Overall, this study provides important insights into the mastication trait formation of Valencia orange fruit.

Development of SNP molecular markers of Pinus bungeana based on SLAF-seq technology

Development of SNP molecular markers of Pinus bungeana based on SLAF-seq technology

Genetic diversity and evolutionary patterns of Taraxacum kok-saghyz Rodin.

Taraxacum kok‐saghyz Rodin (TKS) is an important potential alternative source of natural inulin and rubber production, which has great significance for the production of industrial products. In this study, we sequenced 58 wild TKS individuals collected from four different geography regions worldwide to elucidate the population structure, genetic diversity, and the patterns of evolution. Also, the first flowering time, crown diameter, morphological characteristics of leaf, and scape of all TKS individuals were measured and evaluated statistically. Phylogenetic analysis based on SNPs and cluster analysis based on agronomic traits showed that all 58 TKS individuals could be roughly divided into three distinct groups: (a) Zhaosu County in Xinjiang (population AB, including a few individuals from population C and D); (b) Tekes County in Xinjiang (population C); and (c) Tuzkol lake in Kazakhstan (population D). Population D exhibited a closer genetic relationship with population C compared with population AB. Genetic diversity analysis further revealed that population expansion from C and D to AB occurred, as well as gene flow between them. Additionally, some natural selection regions were identified in AB population. Function annotation of candidate genes identified in these regions revealed that they mainly participated in biological regulation processes, such as transporter activity, structural molecule activity, and molecular function regulator. We speculated that the genes identified in selective sweep regions may contribute to TKS adaptation to the Yili River Valley of Xinjiang. In general, this study provides new insights in clarifying population structure and genetic diversity analysis of TKS using SNP molecular markers and agronomic traits.

Mathematical modeling for genomic selection in Serbian dairy cattle

This manuscript has come as a result of an efficient breeding program in Serbian cattle populations for some economically important traits. Genomic selection in the last two decades has been the main challenge in animal breeding programs and genetics. Many SNP markers are used in statistical analysis in predicting the accuracy of breeding values for young animals without their performance. The new breeding tendency in the selection of young animals allows their genetic progress with reducing cost. In this study, 92 Holstein cows from various regions in Serbia were analyzed based on SNP molecular markers. Within this investigation, an empirical model was developed for the prediction of Yield Traits and Fertility Traits variables, according to Key traits data for dairy cattle. The developed model gave a reasonable fit to the data and successfully predicted Yield Traits (such as Fat and Protein Percent, Cheese Merit, Fluid Merit, and Cow Livability) and Fertility Traits variables (such as Sire Calving Ease, Heifer Conception Rate, Cow Conception Rate, Daughter Stillbirth, Sire Stillbirth, and Gestation Length). A total of 92 dairy cattle data were used to build a prediction model for the prediction of Yield Traits and Fertility Traits variables. The artificial neural network model, based on the Broyden- Fletcher-Goldfarb-Shanno iterative algorithm, showed good prediction capabilities (the r2 values during the training cycle for the before mentioned output variables were in the range between 0.444 and 0.989).

Análisis genómico de diversidad y estructura genómica de las poblaciones bovinas de la raza mexicana de Lidia

La raza bovina de Lidia ha sido seleccionada desde el siglo XIII para participar en festejos sociales reconocidos bajo el término de “Tauromaquia”. En la actualidad forman parte de la identidad de las culturas regionales de varios países. En México, la raza se especializó a finales del siglo XIX cuando cuatro familias mexicanas importaron un número reducido de bovinos de España. De estas importaciones actualmente solo permanecen las líneas derivadas de las familias Llaguno y González. En la familia Llaguno se llevaron diferentes estrategias de reproducción. Antonio Llaguno cruzó los recién importados bovinos españoles entre sí; de dichas cruzas derivó la línea que actualmente es reconocida como “Pura”. Por otro lado, Julián Llaguno realizó cruzas entre hembras criollas con machos españoles, línea conocida como “Impura”. Por último, entre1996 y 1997, un grupo de ganaderos importó bovinos pertenecientes a ciertos encastes españoles como Domecq, Murube, Santa Coloma, entre otros. El objetivo del presente estudio fue investigar la diversidad genómica, estructura poblacional, niveles de endogamia y relaciones genéticas entre las poblaciones de Lidia mexicana, utilizando marcadores moleculares de tipo SNP. La población fue dividida en cinco grupos: Antonio Llaguno, Julián Llaguno, González, y dos grupos que incluían bien importaciones recientes de origen Domecq, o bien importaciones recientes de origen Santa Coloma. Los resultados permiten apreciar diferentes orígenes genéticos dentro de la familia Llaguno en función de su origen histórico: Antonio y Julián. En el resto de grupos también se observa una clara diferenciación genética. Este aislamiento genético entre poblaciones de Lidia mexicana es una característica de su singularidad.

INTROGRESIÓN DE LOS GENES DE RESISTENCIA A ROYA AMARILLA Yr5a Y Yr15 EN EL CULTIVAR DE TRIGO HARINERO BORLAUG 100

La producción de trigo (Triticum aestivum L.) es afectada por diversos factores bióticos como la roya amarilla causada por Puccinia striiformis f. sp. tritici. Este patógeno tiene el potencial de ocasionar pérdidas en el rendimiento de hasta 100 % si los cultivares susceptibles son infectados en etapas tempranas de crecimiento. La mejor estrategia para el manejo de esta enfermedad es el uso de genotipos resistentes. El objetivo del presente estudio fue transferir los genes Yr5a y Yr15 de Blanca Grande 515 al cultivar Borlaug 100 (parental recurrente) y obtener líneas resistentes manteniendo las características agronómicas de la misma. Se obtuvieron 389 líneas a partir de dos y tres retrocruzas entre Borlaug 100 y Blanca Grande 515, las cuales se evaluaron en plántula y en planta adulta en campo contra el aislado CMEX14.25. La presencia de los genes se corroboró con marcadores moleculares SNP. Los resultados indicaron que Yr5a, previamente reportado en estado homocigótico en Blanca Grande 515, en realidad se encuentra en forma heterocigótica, por lo que el número de líneas positivas a este gen fue bajo en comparación con las positivas para Yr15. El número de combinaciones de genes encontradas fue menor que lo esperado y sólo se encontraron líneas positivas para los dos genes de interés después de realizar la segunda retrocruza. Del total de líneas generadas, se identificaron tres que poseen los genes Yr5a y Yr15 en condición homocigótica, una semejante a Blanca Grande 515 y otra con ambos genes en condición heterocigótica. Dos de las tres primeras líneas también poseen el gen Yr17, proveniente de Borlaug 100, y fenotípicamente fueron muy semejantes a ese cultivar, lo que demuestra que el método de retrocruzamiento es efectivo para transferir y acumular mayor cantidad de genes para lograr resistencia durable manteniendo el fenotipo del progenitor recurrente.

Morphological and molecular description of three commercial Capsicum varieties: a look at the correlation of traits and genetic distancing

Capsicum L. is a genus of vegetables with a high global demand due to the pungency of its fruits. The species C. annuum L., C. frutescens L. and C. chinense Jacq. are the most cultivated and are closely related, belonging a group known as the annuum complex. Within these species, there are varieties with great morphological diversity that are widely exploited commercially. In this study, morphological measurements were performed on commercial chili pepper varieties, including Tabasco (C. frutescens), Cayenne (C. annuum) and Habanero (C. chinense), which are widely cultivated in the southwestern Colombian region, to generate a detailed phenotypic description and determine the correlation between traits. Additionally, microsatellite and SNP molecular markers were implemented to evaluate the genetic distances between them. The probability of forming hybrids between these varieties was also evaluated. Descriptive statistical parameters were estimated for the traits evaluated in forty plants of each commercial variety, and positive morphological correlations were found between the number of seeds, fruit mass and fruit length, as well as the independence or low correlation of this trait group versus the fruit diameter, day of first flower, number of fruits and productivity. The fruit capsaicin contents were estimated, with Habanero being the most pungent with 54.37 ± 5.83 mg/g. Molecular characterization using microsatellite markers and SNPs demonstrated the absence of heterozygous individuals and wide genetic distances between the commercial varieties evaluated; this outcome supported the impossibility of forming hybrids. The high genetic similarity among individuals within varieties could be useful to explore phenotypic plasticity in different environments. The methodology used here proved to be robust in testing trait correlation and cultivar genetic distancing, showing a look at the morphological and molecular relationships inside the genus which can be improved with the inclusion of more varieties. This information is especially useful for growers and breeders who wish to use and evaluate these plant materials.

Development and application of two novel functional molecular markers of BADH2 in rice

Fragrance is one of the most important quality traits in rice, and the phenotype is attributed to the loss-of-function betaine aldehyde dehydrogenase ( BADH2 ) gene. At least 12 allelic variations of BADH2 have been identified, and some of these have been applied to rice fragrance breeding using traditional molecular markers and Sanger sequencing techniques. However, these traditional methods have several limitations, such as being very expensive, imprecise, inefficient, and having security issues. Thus, a new molecular marker technology must be developed to improve rice fragrance breeding. In this study, more than 95% of the cultivated fragrant rice varieties belonged to a 7-bp deletion in exon 2 ( badh2-E2 ) or an 8-bp deletion and 3-bp variation in exon 7 ( badh2-E7 ). Both allelic variations resulted in the loss of function of the badh2 gene. We developed two novel SNP molecular markers, SNP_badh2-E2 and SNP_badh2-E7, related to the alleles. Their genotype and phenotype were highly cosegregated in the natural variation of rice accessions, with 160 of the 164 fragrant rice varieties detected with the two markers. These markers cosegregated with the fragrance phenotype in the F 2 population. Two functional SNP molecular markers of badh2-E2 and badh2-E7 allelic variations were developed. These functional SNP molecular markers can be used for genotype and genetic improvement of rice fragrance through marker-assisted selection and will significantly improve the efficiency of fragrant rice breeding and promote commercial molecular breeding of rice in the future. How to cite: Li W, Zeng X, Li S, et al. Development and application of two novel functional molecular markers of BADH2 in rice. Electron J Biotechnol 2020;46. https://doi.org/10.1016/j.ejbt.2020.04.004 .