Expression levels of three bacterial blight resistance genes against K3a race of Korea by molecular and phenotype analysis in japonica rice (O. sativa L.)

Jung-Pil Suh,Tae-Hwan Noh,Ki-Young Kim,Jeong-Ju Kim,Yeon-Gyu Kim,Kshirod K. Jena 한국작물학회 2009 Journal of crop science and biotechnology Vol.12 No.3

Bacterial blight, caused by Xanthomonas oryzae pv. oryzae (Xoo) is a destructive disease of rice in the major rice growing countries of Asia. In 2003, a serious bacterial blight epidemic occurred in the southwestern coastal areas in Korea, causing significant yield loss due to the emergence of a new race, K3a. IR24 near-isogenic lines containing Xa4, xa5, Xa7 and Xa21 genes conferred different degrees of resistance to the most virulent K3a isolate, HB01009 in an inoculation experiment in the greenhouse. Expression levels of the resistance genes, Xa4, xa5 and Xa21 were studied in two F2 populations derived from the crosses between elite japonica cultivars and an advanced backcross breeding line possessing Xa4, xa5 and Xa21 genes. F2 progenies segregated for K3a resistance (R) and susceptible (S) phenotypes in a ratio of 3(R):1(S) indicated that K3a resistance was controlled by a major dominant gene. Three PCR markers tightly linked to the resistance genes Xa4, xa5 and Xa21 confirmed the presence of the genes and their interaction with each gene. This study demonstrated that the Xa21 gene dominantly contributed to K3a resistance. However, the Xa4 gene also contributed to the full expression of resistance. The level of expression of strong resistance to K3a race was attributed to the presence of Xa21 and Xa4 genes irrespective of the presence of xa5 gene. Our results suggest that the R-gene combinations of Xa4+Xa21 could be a useful and effective strategy toward improving resistance to K3a race of Korean japonica cultivars. Bacterial blight, caused by Xanthomonas oryzae pv. oryzae (Xoo) is a destructive disease of rice in the major rice growing countries of Asia. In 2003, a serious bacterial blight epidemic occurred in the southwestern coastal areas in Korea, causing significant yield loss due to the emergence of a new race, K3a. IR24 near-isogenic lines containing Xa4, xa5, Xa7 and Xa21 genes conferred different degrees of resistance to the most virulent K3a isolate, HB01009 in an inoculation experiment in the greenhouse. Expression levels of the resistance genes, Xa4, xa5 and Xa21 were studied in two F2 populations derived from the crosses between elite japonica cultivars and an advanced backcross breeding line possessing Xa4, xa5 and Xa21 genes. F2 progenies segregated for K3a resistance (R) and susceptible (S) phenotypes in a ratio of 3(R):1(S) indicated that K3a resistance was controlled by a major dominant gene. Three PCR markers tightly linked to the resistance genes Xa4, xa5 and Xa21 confirmed the presence of the genes and their interaction with each gene. This study demonstrated that the Xa21 gene dominantly contributed to K3a resistance. However, the Xa4 gene also contributed to the full expression of resistance. The level of expression of strong resistance to K3a race was attributed to the presence of Xa21 and Xa4 genes irrespective of the presence of xa5 gene. Our results suggest that the R-gene combinations of Xa4+Xa21 could be a useful and effective strategy toward improving resistance to K3a race of Korean japonica cultivars.

Marker Assisted Selection-A New Paradigm in Plant Breeding

Kshirod K. Jena,Huhn Pal Moon,David J. Mackill 한국육종학회 2003 한국육종학회지 Vol.35 No.3

The production and productivity of major crop plants have reached their plateu during the past decade because of adop-tion of green revolution technologies. However, further increase in production of cereals with improved cereal quality is imperativeto fe

Genetic diversity in Oryza species and use of wild species in rice improvement

Jena Kshirod K.,Brar Darshan S.,Mackill David J. 한국작물학회 2003 한국작물학회 학술발표대회 논문집 Vol.2003 No.-

Effectiveness of marker-assisted backcrossing breeding for biotic resistance in rice

Jung-Pil Suh,Kshirod K. Jena,Young-Chan Cho,Ji-Ung Jeung,Yong-Jae Won,Im-Soo Choi,Jeom-Ho Lee,Myeong-Ki Kim,Chung-Kon Kim 한국육종학회 2013 한국육종학회 심포지엄 Vol.2013 No.07

The transfer of a biotic resistance gene from indica rice cultivars into japonica cultivars by conventional breeding methods often difficult due to high sterility of the progenies, poor plant type, and linkage drag. Molecular markers provide opportunities to map resistance genes and accelerate the application of marker-assisted backcross(MAB) breeding through the precise transfer of target genomic regions into the recurrent parent. The basis of MAB breeding is to transfer a specific gene/allele of the donor parent into the recurrent parent genome while selecting against donor introgressions across the rest of the genome. The effectiveness of MAB breeding depends on the availability of closely linked DNA markers for the target locus, the size of the population, the number of backcrosses and the position and number of markers for background selection. We have successfully developed Bph18 version of the commercially cultivated japonica elite cultivar by using MAB and incorporating the resistance gene Bph18 that conferred enhanced resistance to BPH. MAB breeding provides a new opportunity for the selective transfer of biotic resistance genes into elite indica rice cultivars devoid of linkage drag. In additon, molecular markers precisely estimate the introgression of chromosome segments from donor parents and can speed up the recipient genome recovery via background selection.

A simple, rapid, and high-throughput DNA extraction method for PCR analysis from rice plants

Sung-Ryul Kim,Gynheung An,Kshirod K Jena 한국육종학회 2013 한국육종학회 심포지엄 Vol.2013 No.07

Polymerase chain reaction (PCR) is highly utilized for QTL analysis, positional cloning of valuable genes, and molecular breeding in crop science. Usually those experiments handle DNA samples of many genotypes (up to several thousands). However, many DNA extraction protocols require longer time using harmful chemicals such as chloroform, phenol, and liquid nitrogen. Here, we introduce a new DNA extraction method for PCR with agarose/PAGE analysis from a diversity panel of rice genotypes identified with yield enhancing traits. This protocol consists of four steps including injection of extraction buffer (20 mM Tris-HCl pH9.5, 200 mM KCl, 2 mM EDTA) into the tubes containing leaf tissues and steel balls, and crushing tissues using Geno-Grinder without liquid nitrogen, sample incubation at 65°C, and then centrifugation for removing cell debris. After centrifugation the crude extracts directly used as template DNA for PCR. Through this protocol we could complete F1 hybridity test from approximately 2,100 plants that come from 96 cross combinations with 13 SSR markers. In addition, we tested the DNA quality by PCR amplification of high GC-rich region and large target size (-2kb). From these results our DNA extraction method produces enough DNA quality for PCR and is suitable for large scale molecular analysis from rice plants.

METHOD AND TECHNOLOGY : A Simple DNA Preparation Method for High Quality Polymerase Chain Reaction in Rice

( Sung Ryul Kim ),( Jungil Yang ),( Gynheung An ),( Kshirod K Jena ) 한국육종학회 2016 Plant Breeding and Biotechnology Vol.4 No.1

Preparation of DNA is cumbersome especially in the case of large numbers of plant samples. Several simple plant DNA preparation methods have been developed for use in conjunction with polymerase chain reaction (PCR) analysis. However, those methods have not been adopted widely for rice molecular analysis. We present a new, simple, and inexpensive method using tris-phosphate (TPE) ethylenediaminetetraacetic acid (EDTA) buffer (100 mM tris-HCl pH9.5, 1 M KCl, 10 mM EDTA pH 8.0) without phenol-chloroform extraction and DNA precipitation steps. The method consists of five steps: leaf tissue grinding, incubating in TPE buffer at 65oC for 20 to 90 minutes, diluting extracts with water, centrifuging to sediment tissue debris, and transferring the supernatant for direct use in PCR or storage. Agarose gel analysis of the crude extracts indicated that the method produced intact genomic DNA (gDNA) from young and old leaves of both young seedlings and mature plants. Leaf sample size (0.5 to 8.0 cm long) for DNA preparation was less sensitive to PCR than the previous methods. DNA quality was tested through PCR amplification of various GC content regions and product sizes, and we obtained bands from all samples, indicating that the method produced suitable DNA quality for PCR. gDNAs were stable for longer than eight months at 4oC. This protocol enabled one person to handle several hundred samples in a day and was tested through various PCR-gel analyses such as genotyping of rice T-DNA mutant lines, positional cloning of rice mutant, and high throughput marker-assisted breeding using allele-specific SNP/Indel markers.

Cytological Characterization of Interspecific Hybrids in Rice (Oryza sativa L.)

Eung Gi Jeong,Darshan S. Brar,Kyung Ho Kang,Heung Goo Hwang,Kshirod K. Jena,Ho Yeong Kim,Sang Nag Ahn,Gihwan Yi,Min Hee Nam 한국육종학회 2005 한국육종학회지 Vol.37 No.1

The wild species of Oryza are an important source of useful genes for resistance to biotic and abiotic stresses in rice. In this study, wide hybridization was used to widen the genepool of japonica rice cultivars. Interspecific crosses were made between three japonica rice cultivars and four wild species of the genus Oryza. Hybrids were produced normally through crosses between O. sativa (AA genome) and O. rufipogon (AA genome). However, hybrids between O. sativa and other distantly related wild species such as O. officinalis (CC genome), O. minuta (BBCC genome), and O. alta (CCDD genome) were produced following hybridization and embryo rescue. The hybrids were intermediate in their morphological characters, compared with the parents of cultivated and wild species. The hybrids between O. sativa and O. rufipogon were partially sterile but the hybrids between O. sativa and O. officinalis, O. sativa and O. minuta, O. sativa and O. alta were completely sterile. Cytological analysis of the interspecific hybrids, AA x CC, AA x BBCC, AA x CCDD showed irregular meiosis with predominant occurrence of univalents and unequal distribution of chromosomes at anaphase 1 of meiosis. Genomic In-situ Hybridization (GISH) analysis showed the presence of O. minuta chromosomes in the F₁ hybrids between O. sativa and O. minuta cross.

Molecular markers from QTLs fine mapped regions of the candidate genes calmodulin-binding transcription activator (CMTA) and integral membrane Duf6 domain containing protein are associated with seedling cold tolerance in rice (Oryza sativa L.)

Suk-Man Kim,Jung-Pil Suh,Chung-Koon Lee,Yeong-Gyu Kim,Kshirod K. Jena 한국육종학회 2012 한국육종학회 심포지엄 Vol.2012 No.07

Cold stress at the seedling stage is a major threat to rice production. Cold tolerance is controlled by complex genetic factors. We used an F7 recombinant inbred line (RIL) population of 123 individuals derived from the cross of a cold-tolerant japonica and a cold-sensitive indica cultivars, for QTL mapping. Phenotypic evaluation of the parents and RILs in an 18/8oC (day/night) cold-stress regime showed continuous variations for cold tolerance or sensitivity. Six QTLs for seedling cold tolerance were identified on chromosomes 1, 2, 4, 10, and 11 with percent phenotypic variation (R2) ranging from 6.1% to 16.5%. Three main-effect QTLs (qSCT1, qSCT4, and qSCT11) were detected in all cold-tolerant RILs which explained high sum of phenotypic variation (SPV) ranging from 27.1% to 50.6%. Two QTLs (qSCT1 and qSCT11) on chromosomes 1 and 11 were fine mapped. The marker In1-c3 from ORF LOC_Os01g69910 of the BAC clone B1455F06 encoding calmodulin-binding transcription activator (CAMTA) and another marker, In11-d1 from ORF LOC_Os11g37720 (Duf6 gene) of the BAC clone OSJNBa0029K08, co-segregated with seedling cold tolerance. These two InDel markers amplified 241-bp and 158-bp alleles, respectively, in cold-tolerant RILs, and in the cold-tolerant donor Jinbu, which were absent in cold-sensitive parent BR29 and cold-sensitive RILs.

Development of elite japonica rice breeding lines for BPH resistance by marker-assisted backcross breeding

Jung-Pil Suh,Ji-Ung Jeung,Jong-Hee Lee,Jeom-Ho Lee,Sae-Jun Yang,Yeon-Gyu Kin,Kshirod K. Jena 한국작물학회 2008 한국작물학회 학술발표대회 논문집 Vol.2008 No.10

Brown planthopper (BPH) is a major insect pest of tropical indica and temperate japonica rice in Asia and Africa. A major BPH resistance gene, Bph18 derived from IR65482-7-216-1-2 has been fine mapped on chromosome 12 and confers strong resistance to the Korean biotype of BPH. The Bph18 gene is tightly linked to the STS marker, 7312.T4A and is non-allelic to previously reported resistance genes present on chromosome 12. The Bph18 gene has been transferred into two elite japonica cultivars (Jinbubyeo and Junambyeo) background through marker-assisted backcross breeding (MAB) strategy. Foreground selection using STS markrs linked to the Bph18 gene in advanced backcross progenies confirmed homozygous marker alleles associated with BPH resistance. Background selection of the breeding lines with 260 simple repeat (SSR) markers revealed rapid conversion toward recurrent parent genotypes with less donor chromosomal segments (5.3-16.7%). Major agronomic traits of the progenies were analyzed and some breeding lines have agronomic traits comparable to the recurrent parent. One breeding line (S.523) with multiple-resistance to BPH and major diseases, desirable agronomic traits and grain quality has been recommended for regional testing in Korea. MAB is the suitable strategy to incorporate new genes into susceptible japonica to develop elite breeding lines.

Identification and fine-mapping of a new resistance gene, Xa40, conferring resistance to bacterial blight races in rice (Oryza sativa L.)

Kim, Suk-Man,Suh, Jung-Pil,Qin, Yang,Noh, Tae-Hwan,Reinke, Russell F.,Jena, Kshirod K. Springer-Verlag 2015 Theoretical and applied genetics Vol.128 No.10