Mapping a Rice Glabrous Gene Using Simple Sequence Repeat Markers

Inheritance analysis and gene mapping of the glabrous trait were conducted using the crosses between a pubescent rice material Chuanxiang 29B （an aromatic elite maintainer line of hybrid rice）, and two glabrous rice materials, Lemont and R2 （R2 is a new restorer line of hybrid rice）. All F1 plants from the two crosses had pubescent leaves, confirming that the pubescence trait in Chuanxiang 29B is dominant over the glabrous trait. In F2 individuals, the segregation ratio of three pubescent to one glabrous plant suggests that a single glabrous gene was present in Lemont and R2. Using the bulked segregant analysis, the linkage analysis of the simple sequence repeat markers showed that the glabrous gene, gl-1, was flanked by GL311 and GL8 with the genetic distances of 0.19 and 0.92 cM on chromosome 5, respectively.

cDNA Cloning and Sequence Analysis of Rice Sbe1 and Sbe3 Genes

Two starch-branching enzyme (SBE) in rice, is known to be a key enzyme in amylopectin biosynthesis. The cDNA of two SBE(starch-branching enzyme) genes Sbe1 and Sbc3 encoding SBE Ⅰ and SBE Ⅲ (two major isoforms in rice) were cloned by an improved RT-PCR technique, from a template cDNA library derived from the total mRNAs extracted from the immature seeds of a japonica rice Wuyunjing 7. DNA sequence analysis showed that the size of the cloned Sbe1 and Sbe3 cDNAs were 2490 and 2481 bp long, respectively, including their entire coding sequences. Comparison analysis indicated that the nucleotide sequence of Sbe3 was the same as that of sbc3 (Genbank Accession No. D16201) as reported previously. There were only four base-pairs difference, which resulted in changes of two deduced amino acids between the cloned Sbel cDNA and the reported sbe1 (Genbank Accession No. D11082). The cloned Sbe1 and Sbe3 cDNAs make it possible to improve rice starch quality through genetic engineering

A Cleaved Amplified Polymorphic Sequence Marker to Detect Variation in Wx Locus Conditioning Translucent Endosperm in Rice

The translucent endosperm trait in a japonica rice variety ＇Kantou 194＇ is controlled by a Wx-mq gene which is allelic to Wx locus by genetic analysis and allelic test. The amylose content analysis showed that an intermediate amylose content between those of glutinous and non-glutinous rice existed in endosperm of homozygous Wx-mq genotype. The slight changes of amylose content in different varieties and F1 grains with an identical Wx-mq genotype might be influenced by dissimilar genetic background. To identify the Wx-mq genotype simply and rapidly, a cleaved amplified polymorphic sequence （CAPS） marker was designed. The result from the molecular detection indicated that it could be used for marker-assisted selection for low amylose content varieties in rice breeding.

Amplification and sequencing of a sulfur-rich 10kd prolamin gene from rice seeds

Nutritious value of seed storage protein is low due to deficiency in essential amino acid contents. Cereals are mainly deficient in lysine and legumes in sulfur-containing amino acids (methionine and cysteine). So far, several sufur-rich seed protein genes have been isolated and the essential amino acid contents of seed proteins were increased in transgenic tobacco and Brassica napus. In this paper we report the isolation and sequencing of the 10kd prolamin gene from seeds. Poly(A) RNA were prepared from the immature endosperms of japonica rice, Sachiminori, 10 d after flowering. Complementary DNAs were synthesized according to Promega Instruction Manual. Two primers were synthesized and their sequences were Primer Ⅰ: CGTCTACACCATCTGGAATC, Primer Ⅱ: GTGTTTGCAGATAGTATGC. The amplified fragraents were inserted into the Sma I site of pGEM-7zf(+) and was used to transform E. coli JM101 after PCR reaction. DNA sequence were determined by Sanger’s Chain-termination method. Synthesis of cDNA. Using mRNAs as

Mapping of a New Gene Wbph6(t) Resistant to the Whitebacked Planthopper, Sogatella furcifera, in Rice

A rice population consisting of 90 TN1/Guiyigu F3 lines was employed to analyze the linkage between DNA markers and a new gene Wbph6(t) conferring resistance to whitebacked planthopper, Sogatella furcifera By using the mapping approach of bulked extremes and recessive class, Wbph6(t) was mapped onto the short arm of chromosome 11 with a genetic distance of 21.2 cM to SSLP marker RM167.

Genetic Polymorphism of Wx Gene and Its Correlation with Main Grain Quality Characteristics in Rice

The allelic variation of the Wx gene in 50 non-glutinous rice varieties （lines） was analyzed by using the microsatellite marker RM190 [for （CT）n simple sequence repeat （SSR）] and cleaved amplified polymorphic sequence（CAPS） marker 484/W2R-ACCⅠ[for G/T single nucleotide polymorphism （SNP）]. Six homozygous （CT）n types, namely （CT）20, （CT）19, （CT）18, （CT）17, （CT）16, （CT）14, （CT）11 and （CT）10, and a heterozygous genotype （CT）11/（CT）18 were detected for RM190, of which （CT）11 and （CT）18 were predominant. Two homozygous Wx genotypes （G/G and T/T） and one heterozygous （G/T） were detected using 484/W2R-ACC Ⅰ. Most of the materials with a RM190 of （CT）11 were G/G for SNP of 484/W2R-ACC Ⅰ, while T/T for SNP was predominantly appeared in materials with （CT）18. The materials tested could be grouped into 10 categories using the two markers together. Results indicated that 59.3% variance of amylose content was attributed to the polymorphism of Wx gene revealed by RM190, while 56.1% and 24.6% of the variances in amylose content and gel consistency were respectively to the polymorphism of Wx gene revealed by 484/W2R-ACC Ⅰ. Furthermore, with both SSR and CAPS markers, 72.4% of the variance in amylose content could be explained. In addition, the application prospects of the two markers in breeding were also discussed.

Identification of candidate genes for drought stress tolerance in rice by the integration of a genetic (QTL) map with the rice genome physical map

Genetic improvement for drought stress tolerance in rice involves the quantitative nature of the trait, which reflects the additive effects of several genetic loci throughout the genome. Yield components and related traits under stressed and well-water conditions were assayed in mapping populations derived from crosses of Azucena×IR64 and Azucena×Bala. To find the candidate rice genes underlying Quantitative Trait Loci (QTL) in these populations, we conducted in silico analysis of a candidate region flanked by the genetic markers RM212 and RM319 on chromosome 1, proximal to the semi-dwarf (sd1) locus. A total of 175 annotated genes were identified from this region. These included 48 genes annotated by functional homology to known genes, 23 pseudogenes, 24 ab initio predicted genes supported by an alignment match to an EST (Expressed sequence tag) of unknown function, and 80 hypothetical genes predicted solely by ab initio means. Among these, 16 candidate genes could potentially be involved in drought stress response.

Comparative mapping of QTLs for Al tolerance in rice and identification of positional Al-induced genes

Aluminum (Al) toxicity is the major factor limiting crop productivity in acid soils. In this study, a recombinant inbreed line (RIL) population derived from a cross between an Al sensitive lowland indica rice variety IR1552 and an Al tolerant upland japonica rice variety Azucena, was used for mapping quantitative trait loci (QTLs) for Al tolerance. Three QTLs for relative root length (RRL) were detected on chromosome 1, 9, 12, respectively, and 1 QTL for root length under Al stress is identical on chromosome 1 after one week and two weeks stress. Comparison of QTLs on chromosome 1 from different studies indicated an identical interval between C86 and RZ801 with gene(s) for Al tolerance. This interval provides an important start point for isolating genes responsible for Al tolerance and understanding the genetic nature of Al tolerance in rice. Four Al induced ESTs located in this interval were screened by reverse Northern analysis and confirmed by Northern analysis. They would be candidate genes for the QTL.

A mitochondrial phosphate transporter,McPht gene,confers an acclimation regulation of the transgenic rice to phosphorus deficiency

Phosphate transporters play an important role in promoting the uptake and transport of phosphate in plants. In this study, the McPht gene from the Mesembryanthemum crystallinum, a mitochondrial phosphate transporter, was isolated and constructed onto a constitutive expression vector carrying 35S：：GFP, and the recombinant constructs were transferred into Oryza sativajaponica L. cv. Kitaake to investigate the regulatory role of the McPhtgene under phosphorus deficiency. The McPhtgene encodes a protein of 357 amino acids with six transmembrane domains and is located to the mitochondria, and the mRNAtranscripts of the McPht gene are highly accumulated in the shoots of M. crystallinum in response to phosphorus deficiency. However, more mRNA transcripts of the McPht gene were accumulated in the roots of the transgenic rice under phosphorus deficiency. Measurements showed that the transgenic rice demonstrated an enhanced promotion in the root development, the root activities, and phosphate uptake under phosphorus deficiency. Transcriptome sequencing showed that the transgenic rice exhibited total of 198 differentially expressed genes. Of these, total of 154 differentially expressed genes were up-regulated and total 44 genes were down-regulated comparing to the wild type in response to phosphorus deficiency. The selective six genes of the up-regulated differentially expressed genes showed an enhanced increase in mRNA transcripts in response to phosphorus deficiency, however, the transcripts of the mitochondrial carrier protein transporter in rice, a homologous gene of the McPht, in both the transgenic line and the wild type had no obvious differences. Functional enrichment analyses revealed that the most of the up-regulated genes are involved in the cytoplasmic membrane-bounded vesicle, and most of the down-regulated genes are involved in the mitochondrion and cytoplasmic membrane-bounded vesicle. The differentially expressed genes were highly enriched in plant secondary metabolisms and plant-pathogen interaction. These results indicated that the overexpression of the McPht gene might participate in the physiological adaptive modulation of the transgenic rice to phosphorus deficiency by up- or down-regulating the differentially expressed genes.

Functional Marker Development and Effect Analysis of Grain Size Gene GW2 in Extreme Grain Size Germplasm in Rice

GW2 is an important gene that regulates grain width and weight. We used cDNA clone to obtain the sequences of GW2 from large- and small-grained rice varieties, TD70 and Kasalath, respectively. Then, we developed a dCAPS （derived cleaved amplified polymorphic sequence） marker on the basis of the sequence difference between functional and nonfunctional GW2 genes to analyze the genotypes and phenotypes of recombinant inbred lines. Results showed that the sequence of GW2To7~ had a single nucleotide deletion at site 316 that generates a termination codon. This codon terminated the GW2 protein in advance. By contrast, the sequence of GW2Kasalath encoded an intact protein. A novel dCAPS marker was designed in accordance with a base A deletion at site 316 of the sequence. After the PCR product was digested by Apol, TD70 showed 21 and 30 bp fragments, and Kasalath showed a 51 bp fragment. Up to 82 lines contained GW2TDTO, and 158 lines contained GW2Kasalath. The lines that contained TD70 alleles displayed substantial increases in width and 1000-grain weight. This result suggested that GW2 played a critical role in rice breeding.

Transgenic japonica rice expressing the cry1C gene is resistant to striped stem borers in Northeast China

Rice production and quality are seriously affected by the lepidopteran pest,striped stem borer(SSB),in Northeast China.In this study,a synthetic cry1 C gene encoding Bacillus thuringiensis(Bt)δ-endotoxin,which is toxic to lepidopteran pest,was transformed into a japonica rice variety(Jigeng 88)in Northeast China by Agrobacterium-mediated transformation.Through molecular detection and the Basta resistance germination assay,a total of 16 single-copy homozygous transgenic lines were obtained from 126 independent transformants expressing cry1 C.Finally,four cry1 C-transgenic lines(JL16,JL23,JL41,and JL42)were selected by evaluation of the Cry1 C protein level,insect-resistance and agronomic traits.The cry1 C-transgenic lines had higher resistance to SSB and higher yield compared with non-transgenic(NT)control plants.T-DNA flanking sequence analysis of the transgenic line JL42 showed that the cry1 C gene was inserted into the intergenic region of chromosome 11,indicating that its insertion may not interfere with the genes near insertion site.In summary,this study developed four cry1 C-transgenic japonica rice lines with high insect resistance and high yield.They can be used as insect-resistant germplasm materials to overcome the problem of rice yield reduction caused by SSB and reduce the use of pesticides in Northeast China.

In silico Cloning and Bioinformatics Analysis of OsARAB-1 Gene from Rice

[ Objective] This study aimed to perform in silico cloning and bioinformatics analysis of OsARAB-1 gene from rice. [Method] Using NP 174188.1 as a query probe, the full-length cDNA sequence of OsARAB-1 gene was obtained by in silico cloning. The nucleotide sequence and protein sequence were analyzed by bioinformatics software. [Result] The full-length cDNA of OsARAB-1 gene was obtained. The coding sequence （CDS） of OsARAB-1 gene is 1 086 bp in length, encoding a protein of 361 amino acid residues. OsARAB-1 gene was located in the genome sequence NC 008398.2 （6 769 813 -6 773 213 bp segment） of rice chro- mosome 5 by in silico mapping. OsARAB-1 protein is an extracellular hydrophilic protein, which is relatively stable, slightly alkaline. The secondary structure of OsARAB-1 protein is mainly composed of or-helices and random coils. OsARAB-1 protein has two functional domains ： SGNH hydrolase-type esterase and GDSL li- pase. There are 21 phosphorylation sites and seven O-[3-GlcNAc glycosylation sites. The putative active-site amino acid residues are Ser34, Glyl07, Asn167, Asp333 and His336, respectively. OsARAB-1 protein has the closest genetic relationship with esterase subtype B4FM12 from maize. The expression of OsARAB-1 gene plays an important role in the development and morphogenesis of rice and is related with rice blast resistance. [ Conclusion] This study laid a solid foundation for cloning and functional identification of OsARAB-1 gene with experimental methods.

籼粳杂种不育的遗传分析和候选基因鉴定

[目的]籼粳亚种间杂交F1的育性问题严重阻碍了亚种间杂种优势的利用,探究籼粳杂种不育的遗传机理,挖掘新的籼粳杂种不育调控基因,为促进籼粳杂种结实率遗传改良提供理论依据。[方法]粳稻品种Sasanishiki和籼稻品种Habataki杂交后,采用单粒传法自交10代,获得包含95个株系的稳定遗传重组自交系(RIL),基于Illumina平台,对双亲和RIL进行高通量测序,在全基因组水平分析RIL中Habataki血缘的分布与比例,进而鉴定偏分离区域作为潜在的籼粳杂种不育位点。同时,剖析“全球3000份水稻核心种质资源重测序计划”中典型籼粳稻基因组变异数据,对群体水平进一步验证并趋近目标育性基因区域。最终通过序列比对锁定籼粳杂种不育候选基因。应用CRISPR基因编辑技术进行定点基因敲除,对目标基因进行功能验证。[结果]Habataki和Sasanishiki的杂交F1在穗数、每穗粒数和千粒重上体现出明显的杂种优势,但其结实率显著降低,I2-KI镜检发现F1花粉育性显著降低。RIL的全基因组高通量测序在第1、3、5、6、7和12染色体上检测到明显的偏分离,即该区域的基因型趋向于籼稻Habataki。通过序列比对,进一步确定已知育性基因Sc、S5和HSA1为第3、6和12染色体上偏分离区域的目标基因。应用CRISPR基因编辑技术敲除Habataki中Sc-Haba-3的多拷贝,成功改善了其与Sasanishiki杂交F1的花粉育性和结实率,说明该基因可独立行使功能。同时,在第1染色体的偏分离区域发现Habataki和Sasanishiki之间存在复杂的结构性变异,Sasanishiki基因组中一段24.7 kb包含4个预测基因的片段在Habataki中被一段64.8 kb包含10个预测基因的片段取代,此结构性变异可能参与籼粳杂种育性的调控。[结论]检测到多个籼粳杂种不育相关位点,应用CRISPR基因编辑技术敲除Habataki中Sc的多拷贝成功改善其杂种F1育性,确定其为水稻亚种间育性改良的目标基因。同时锁定了第1染色体Sd区域的目标基因。

低甲烷排放转基因水稻对土壤微生物群落的影响

低甲烷排放转基因水稻是实现水稻低碳生产的理想材料。土壤微生物驱动了稻田甲烷的产生,低甲烷排放转基因水稻土壤微生物群落组成的变化不仅影响稻田甲烷排放,也关系到土壤微生态系统的稳定性。通过对细菌16S rRNA基因、真菌ITS基因的高通量测序及mcrA、nifH、amoA和nirS等功能基因的荧光定量PCR,分析了低甲烷排放转基因水稻(86R27-3)与野生型水稻(MH86)土壤微生物群落间的差异。结果显示:稻田土壤细菌群落的α-多样性指数在86R27-3与MH86间无明显差异,且仅在水稻分蘖期86R27-3的土壤真菌群落多样性指数Shannon、Simpson及均匀度指数Pielou_e显著高于MH86(P<0.05);β-多样性分析表明土壤细菌或真菌群落组成在86R27-3与MH86间均没有显著差异;但在水稻齐穗期:86R27-3土壤的放线菌门(Actinobacteria)、罗泽真菌门(Rozellomycota)的相对丰度显著高于MH86(P<0.05),而酸杆菌门(Acidibacteria)、子囊菌门(Ascomycota)的相对丰度显著低于MH86(P<0.05);土壤微生物群落功能预测显示,86R27-3土壤氮、硫和锰代谢细菌功能群丰度显著低于MH86(P<0.05),如分蘖期的土壤硝酸盐还原、硝酸盐呼吸、硫代硫酸盐呼吸及硫呼吸,齐穗期和成熟期的好氧亚硝酸盐氧化及成熟期的锰氧化等;与MH86相比,86R27-3的土壤真菌功能群丰度有减有增,如在水稻不同生育期内的其未定义腐生物银耳目、嗜热囊菌科、镰刀菌属及韦斯特氏菌功能群丰度显著降低(P<0.05),而其分蘖期的动物内共生体腐生生物毕赤酵母属和未定义腐生物马勃科功能群丰度显著提高(P<0.05)。定量PCR分析表明86R27-3土壤中的产甲烷细菌mcrA基因丰度显著低于MH86(P<0.05),同时,土壤固氮菌nifH基因、氨氧化细菌amoA基因及反硝化细菌nirS基因的丰度在86R27-3土壤中也显著降低(P<0.05)。综上所述,低甲烷排放转基因水稻(86R27-3)对土壤细菌或真菌的群落组成没有影响,但可引起主要细菌或真菌种类的相对丰度及某些细菌或真菌功能群丰度发生变化,并显著降低了稻田土壤微生物功能基因丰度。

35份水稻骨干亲本氮高效基因分析及功能分子标记开发

氮是影响水稻产量形成最重要的元素之一,其合理和高效利用是农业可持续发展的重要保障。通过靶向测序技术鉴定了35份水稻骨干亲本OsNPF6.1、NRT1.1B、OsNR2和OsGRF4的基因型,并基于PARMS技术(penta-primer amplification refractory mutation system)开发了OsNPF6.1、OsNR2和OsGRF4的荧光功能分子标记。结果表明,荧光功能分子标记检测结果与靶向测序结果一致,并且明确了靶向测序未明确的氮高效基因型。综上所述,开发的氮高效基因功能分子标记为培育氮高效水稻新品种提供了技术支撑。

Cloning and sequence characteristics of the genomic gene of a rice metallothionein

Northern blot analysis showed that a metallothionein gene, ricMT is expressed strongly in the stem of rice with an expression level that could be more than 100-fold stronger than in leaf blades. The results suggest that the 5’ upstream region flanking the coding sequence of the ricMT may contain a fairly strong promoter To elucidatr its regulation and promoter structure, the genomic clones of ricMT were screened out from a rice genomic library and a fragment of about 4084 bp was sequenced. The fragment included a 5’ upstream region of ca. 2970 bp, a transcription region of ca. 690 bp and a 3’ downstream region of ca. 420 bp. Computer analysis of the sequence homology showed that the 5’ upstream region included a putative TATA box, a putative CAAT box, and a typical metal-responsive element TGCGCGCG. The results will promote further understanding of the mechanisms of gene regulation and metal response of plant metallothionein proteins.

Microsatellite-Aided Screening for Fertility Restoration Genes (Rf) Facilitates Hybrid Improvement

DNA markers enabled to determine the chromosomal locations of the two Rf genes(Rf3 and Rf4) in the wild-abortive cytoplasmic male sterility(WA-CMS) system. Four simple sequence repeats(SSRs) RM171, RM258, RM315 and RM443 were used to detect the allelic status with respect to the fertility restoration genes(Rf3 and Rf4) in 300 rice cultivars or breeding lines. The results revealed that out of 300 lines, 90 lines screened had Rf3, 65 lines had Rf4, and 45 lines had Rf3 and Rf4 alleles. Furthermore, 45 lines selected using SSR markers were mated with a CMS line(IR58025A) to analyze their restoring ability. Offspring of all the test lines except HHZ8-SAL9DT1-Y1, HHZ5-SAL9-Y3-1 and IDSA77 exhibited higher pollen and spikelet fertility(> 80%), thus confirming they bear the Rf alleles. The hybrid offspring of ARH12-6-1-1-B-3-1, IR32307-10-3-2-1 and Sahel 329 had the highest pollen fertility(97.39%, 98.30% and 97.10%, respectively) and spikelet fertility(95.10%, 97.07% and 96.10%, respectively).

Differential RNA Editing of Mitochondrial Genes in WA-Cytoplasmic Based Male Sterile Line Pusa 6A, and Its Maintainer and Restorer Lines

RNA editing changes the nucleotides at the transcript level of mitochondrial genes which results in synthesis of functional proteins.This study was designed to find the editing sites which could be implicated in male fertility restoration and to develop editing based markers for differentiation of cytoplasmic male sterility and maintainer lines from each other.DNA and RNA from young panicles were isolated from three-line system of hybrid rice PRH10,wild abortive(WA)cytoplasm based male sterile(A line Pusa 6A),maintainer(B line Pusa 6B)and restorer(R line PRR78)lines.Pusa 6A and PRR78 having the same WA cytoplasm are allo-nuclear and iso-cytpolasmic lines.The genomic and cDNA amplicons for eight mitochondrial genes(18SrRNA,atp6,atp9,cobII,coxI,coxIII,nadI and rps3)were sequenced and compared.Differences in genomic and cDNA sequences were considered as editing.Two hundred and thirty editing sites having base substitution or insertion/deletion were identified with the highest in 18SrRNA(5.74%)and the lowest in coxI(0.60%).The highest editing sites were observed in fertile maintainer Pusa 6B followed by PRR78 and Pusa 6A,of which random five editing sites in five different rice mitochondrial transcripts namely atp9,cobII,coxIII,rps3 and 18SrRNA were chosen and validated through cleaved amplified polymorphism sequence(CAPS)analysis and found to be partially edited in four genes.The identical editing sites of different mitochondrial genes from maintainer and restorer lines might reflect their possible contribution to fertility restoration of sterile WA cytoplasm.

Molecular Cloning of a Thiol Proteinase Inhibitor Gene and Its Expression in E.coli

A cDNA library was constructed with 1.5×10~6 pfu from rice immature seeds,fromwhich a cDNA clone for rice thiol proteinase inhibitor,oryzacystatin(OC),was isolated byscreening with synthesized oligodeoxynucleotide probe,which contained a 309bp open read-ing frame,84bp 5′-end noncoding region and a poly(A)signal AATAAA at the 3′-end fol-lowed by 31Nt poly(A).Then the coding region of OC was amplified and inserted into thedownstream of λP_RP_L promoter for thermal-inducible expression in E.coli.Shifting the cul-ture temperature from 30℃ to 42℃ led to a high level expression of OC,which exhibited adistinct band of 12.0 kDa and accounted for at least 10% of the total soluble proteins fromSDS-PAGE.The papain-inhibitory activity of the expressed OC was further confirmed.

Identification of QTLs for Blast, Bacterial Blight, and Planthopper Resistance Using SNP-Based Linkage Maps from Two Recombinant Inbred Rice Lines

Rice is the most significant global food security. Several biotic factors limit rice production, breeding biotic-resistant rice has, therefore, become an increasingly important goal. Two elite rice lines, IR71033-121-15 (IR71033) and IR57514-PMI-5-B-1-2 (IR57514), provide potential genes for biotic stress resistance traits. In this study, genotyping by sequencing (GBS) for single nucleotide polymorphism (SNP)-based linkage map construction was used to detect quantitative trait loci (QTLs) for blast (BL), bacterial blight (BB), whitebacked planthopper (WBPH), and brown planthopper (BPH) resistance. IR71033 was derived from Oryza minuta and carried BL, BB, WBPH, and BPH resistance QTLs. IR57514 is a well-adapted rainfed lowland line that carries BL and BB resistance QTLs. Two sets of recombinant inbred line (RIL) populations derived from crosses of KDML105 × IR71033 and KDML105 × IR57514 were used to dissect the genetic basis of disease and insect pest resistance. The RIL populations were evaluated for BL, BB, WBPH, and BPH resistance from 2016 to 2018 at four rice research centers in Thailand. From these, we identified a large number of SNPs through GBS and constructed high-resolution linkage maps. By combining phenotypic evaluation with the GBS data, a total of 24 QTLs on four chromosomes were detected that confered pest resistance and explained 7.3% - 61.4% of the phenotypic variance. These findings should facilitate identifying novel resistance genes and applying marker-assisted selection for resistance to the four major rice pests investigated here. These strategies will improve the resilience and reliability of rice varieties adapted to the low-yielding environment of rainfed lowland areas worldwide.