The cotton aphid, Aphis gossypii (Glover), is one of the most serious pest in seed potato and various vegetable cultivation. The imidacloprid-resistant strain (IR) was over 200-fold more resistant to imidacloprid compared to a susceptible strain (S) as judged by LC50 values. The IR showed cross resistances to other neonicotinoid insecticides. IEF and 2DE analyses revealed that general esterase isozyme patterns in IR were almost identical to those of S. Nevertheless, a significantly overexpressed protein spot was detected in IR. To identify differentially expressed genes in IR, comparative transcriptome analyses based on GS-FLX were conducted using total RNAs extracted from both IR and S strains, which generated ca. 290 Mb reads for each strain. Generally, most nicotinic acetylcholine receptor subunit genes, such as alpha 2 and beta 1, were more transcribed in S than in IR. In contrast, only alpha 5 subunit gene was 1.8 fold more expressed in IR. Seven ATP-binding cassette (ABC) transporter genes were newly identified in A. gossypii, among which only ABCC9 gene was highly expressed in IR. Therefore, this ABCC subfamily, a member of the MRP subfamily which is involved in multi-drug resistance, could be one of the main factors associated with imidacloprid resistance in A. gossypii.

The cotton aphid, Aphis gossypii (Glover), is one of the most serious pest in seed potato and various vegetable cultivation. The imidacloprid resistant strain (IR) was over 200 fold more resistant to imidacloprid compared to the susceptible strain (S) as judged by the LC50 values and IR showed cross resistant to acetamiprid, thiamethoxam, thiacloprid, clothianidin. By using the suppression subtractive hybridization method, a imidacloprid resistant associated cDNA library was constructed in adult cotton aphid. In total 115 differentially expressed cDNA clones were obtained. Any point mutation detected in nicotinic acetylcholine receptor alpha 1~5 and beta 1 subunits in the IR. Based on IEF, the IR general esterase isozyme banding patterns were identical with that of S.

서울시내에서 유통되는 식품과 식품접객업소(집단급식소 포함)의 조리식품을 대상으로 식중독 원인균 분석 및 위생미생물 검사를 실시한 결과, 분리된 대장균의 항생제 감수성 시험을 통하여 이들의 내성 정도를 파악하고, 내성 유전자와 병원성유전자의 분포도 알아보았다. 모두 1313건 의 샘플 중 50건에서 대장균이 검출되어 3.8%의 검출률을 보였다. 이중 육회 1건에서 장출혈성대장균 O26 1건, 김밥 에서 장병원성대장균 1건이 각각 검출되었다. 50건의 대장 균중 50%가 16종의 항생제에 모두 감수성을 보였으며 내성이 높게 나타난 항생제는 ampicillin(36%), amoxicillin/ clavulanic acid(32%) 그리고 tetracycline(22%)의 순이었다. 이들의 내성유전자 분포는 TEM이 1건, tetB 4건이 각각 검출되었다.

The stable fly, Stomoxys calcitrans L., is an important pest of livestock. Stable flies are considered as mechanical vectors of veterinary disease. Pyrethroids and organophosphates have been widely used for stable fly control. To establish resistance monitoring molecular tool, we isolated the partial cDNA and genomic fragments of voltage-sensitive sodium channel and acetylcholinesterase genes encompassing the well known conserved sites for resistance-associated mutations. To examine the current status of stable fly resistance to pyrethroids and organophosphates mediated by the nerve insensitivity mechanism in Korean population of S. calcitrans, DNA-based genotyping in conjunction with residual contact vial (RCV) bioassay were conducted with 11 representative regional field populations. No resistance-associated mutations were detected in these S. calcitrans populations, suggesting that these populations are likely still susceptible to both pyrethroids and organophosphates. Establishment of RCV bioassay protocol and availalbility of target site sequence information will greatly facilitate resistance monitoring of S. calcitrans in the field.

Rice blast, caused by a fungus Magnaporthe oryzae, is one of the most devastating diseases of rice worldwide. Analyzing the valuable genetic resources is important in making progress towards blast resistance. Molecular screening of major rice blast resistance (R) genes was determined in 2,509 accessions of rice germplasm from different geographic regions of Asia and Europe using PCR based markers which showed linkage to twelve major blast R genes, Pik-p, Pi39, Pit, Pik-m, Pi-d(t)2, Pii, Pib, Pik, Pita, Pita/Pita-2, Pi5, and Piz-t. Out of 2,509 accessions, only two accessions had maximum nine blast resistance genes followed by eighteen accessions each with eight R genes. The polygenic combination of three genes was possessed by maximum number of accessions (824), while among others 48 accessions possessed seven genes, 119 accessions had six genes, 267 accessions had five genes, 487 accessions had four genes, 646 accessions had two genes, and 98 accessions had single R gene. The Pik-p gene appeared to be omnipresent and was detected in all germplasm. Furthermore, principal component analysis (PCA) indicated that Pita, Pita/Pita-2, Pi-d(t)2, Pib and Pit were the major genes responsible for resistance in the germplasm. The present investigation revealed that a set of 68 elite germplasm accessions would have a competitive edge over the current resistance donors being utilized in the breeding programs. Overall, these results might be useful to identify and incorporate the resistance genes from germplasm into elite cultivars through marker assisted selection in rice breeding.

Four bacterial blight resistance genes, Xa1+Xa3+xa5+Xa21, pyramid elite japonica rice lines were developed for enhancing the resistance of rice against Xanthomonas oryzae pv. oryzae in Korea. Seven doubled haploid (RDL1-7) and ten F6 lines (RPL1-10) having Xa1+Xa3+xa5+Xa21 which were derived from the cross between Ilmi, high grain quality japonica rice cultivar carrying Xa1, and Iksan575, elite line carrying Xa3+xa5+Xa21, were developed using marker-assisted selection for resistance genes and phenotypic selection for bacterial blight resistance and agronomic traits. Among resistance genes combinations in F2 population, four resistance genes combination, Xa1+Xa3+xa5+Xa21, showed the highest resistance and conferred the enhanced resistance than three genes combination, Xa3+xa5+Xa21. Four genes pyramid lines (RDL and RPL) showed broad-spectrum resistant against 16 Korean bacterial blight isolates and the yield and quality of the lines did not alter by the inoculation of K3a, the most virulent race in Korea. In addition, these lines had excellent plant type and exhibited more enhanced yield than previously developed resistant cultivars. Four bacterial blight resistance genes combination, Xa1+Xa3+xa5+Xa21, was efficient and promising combination and developed lines with four genes could be useful materials and will be applied to the breeding programs for enhancing the resistance of japonica rice against bacterial blight.

This experiments were carried out to know the response to Brown Planthopper(BPH) resistance genes at rice seedling stage using Biotype 1 for develoment of resistant cultivars. Varieties with Bph1, Bph3 and Bph18 genes showed a very strong resistance response, Bph2, Bph6, bph7 and Bph9 genes exhibited moderate resistance. bph5 and bph8 gene retention varieties and Nampyeongbyeo showed a very weak sensitivity in response to BPH. After 72 hours, Nampyeong(no gene) and IR72(Bph3 gene) were showed a feed-preference 690% and 0%, respectively. Results of Antixenosis and seedling resistance response to BPH were grouped into similar by specific resistance genes. Ten days after inoculation, BPH survival rate of vareities with resistance genes were below 30%, whereas Nampyeongbyeo was more than 70%. The results showed that Bph3 and Bph18 genes are highly resistant response against BPH, these genes are very useful for improve the rice cultivars with various resistance genes

Brown planthopper (BPH) is a phloem sap-sucking insect pest of rice, which causes severe yield loss annually. Gayabyeo, a Tongil type rice variety, is known to have broad spectrum resistance to BPH. Before, it was estimated that Gayabyeo has at least two BPH resistance genes. We started a research for mapping resistance genes of Gayabyeo. We did a cross between Taebaekbyeo, a BPH susceptible Tongil type rice variety, and Gayabyeo, We grew F1 plants in winter season of 2014-2015, and planted F2 population in this year. About 100 DNA markers (SSR and InDel markers) showing polymorphism between Gayabyeo and Tabaekbyeo were selected. In addition, we are going to do resequencing Gayabyeo and Taebaekbyeo using Illumina Hiseq2000 to find much more DNA polymorphisms between the two varieties and develop new markers for mapping. The BPH response data will be acquired using F3 plants from the cross between Gayabyeo and Taebaekbyeo next year. In a while, crosses between Gayabyeo and high quality japonica rice varieties are being carried out to introduce BPH resistance genes of Gayabyeo into japonica high quality rice varieties. We expect to develop new DNA markers for BPH resistance genes of Gayabyeo through mapping and produce several japonica high quality rice lines harboring those genes at the end of this project.

Bacterial wilt (BW) caused by Ralstonia solanacearum is one of the most common soil-borne vascular diseases of many solanaceous crops such as pepper and tomato. This study aimed to develop molecular markers closely linked to bacterial wilt resistance genes using a 150 F8 recombinant inbred line (RIL) population obtained from a cross of ‘YCM334’ x ‘Taean’. For pathogen inoculations, R. solanacearum isolate WR-1 was cultured on NB medium at 28℃ for 48 h and a bacterial suspension was adjusted to 1 x 107 to 1 X 108 CFU/mL (A 600 = 0.3 to 0.4). Each RIL and the parents were sown in a 72-cell plastic tray filled with sterilized soil, and the seedlings were inoculated at the 6 to 8 leaf stage using soil-drenching (3 to 5 ml/ plant) inoculation methods with 3 replications. After 10 days post inoculation (dpi), each line was evaluated visually for occurrence of bacterial wilt ranging from 1 (most resistant) to 5 (most susceptible). Two candidate R-response genes, AT4G14130 and AT3G23730, were selected to find SNPs between YCM334 and Taean. In previous transcriptome analysis, these two genes were reported as significantly differentially expressed in Capsicum annuum L. root inoculated with R. solanacearum, which were up-regulated in a resistant genotype. Once the synteny of the gene locations between Arabidopsis and pepper was documented, the sequences on pepper chromosome 12 were obtained from pepper. v.1.55 (http://solgenomics.net). SNP markers associated with resistance to BW will be mapped using pepper RIL population.

MicroRNAs (miRNAs) are a class of non-coding RNAs approximately 21-nt in length which play important roles in regulating gene expression in plants. Although many miRNA studies have focused on a few model plants, miRNAs and their target genes remain largely unknown in hot pepper (Capsicum annuum), one of the most important crops cultivated worldwide. We here employed high-throughput small-RNA and degradome sequencing to comprehensively identify small-RNAs and their targets in pepper. From these, we identified several novel targets of miRNAs, including the major de novo methylation enzyme involved in RNA-directed DNA methylation in plants. Furthermore, we identified several highly abundant 22-nt miRNA families that target conserved domains in NB-LRRs and trigger the production of phased secondary siRNAs. We showed that transient co-expression of can-miR482 with Rpi-blb1, one of the potato NB-LRRs, resulted in the attenuation of the hypersenstive responses in Nicotiana benthamiana, suggesting that interaction between miR-482 family and disease resistance proteins is likely to serve as a conserved trigger for defense mechanism in Solanaceae. This work provides the first reliable draft of the pepper small RNA transcriptome that offers an expanded picture of miRNAs in relation to NB-LRR regulation, providing a basis for understanding the functional roles of miRNAs in disease resistance pepper.

MicroRNAs (miRNAs) are a class of non-coding RNAs approximately 21-nt in length which play important roles in regulating gene expression in plants. Although many miRNA studies have focused on a few model plants, miRNAs and their target genes remain largely unknown in hot pepper (Capsicum annuum), one of the most important crops cultivated worldwide. We here employed high-throughput sequencing to comprehensively identify small RNAs and their targets in pepper. From these, we identified several novel targets of miRNAs, including the major de novo methylation enzyme involved in RNA-directed DNA methylation in plants. Furthermore, we identified several highly abundant 22-nt miRNA families that target conserved domains in NB-LRRs. We showed that transient co-expression of the miRNA with NB-LRRs, resulted in the attenuation of the hypersenstive responses in Nicotiana benthamiana, suggesting that interaction between miRNA family and disease resistance proteins is likely to serve as a conserved trigger for defense mechanism in Solanaceae. This work provides the first reliable draft of the small RNA transcriptome in pepper that offers an expanded picture of miRNAs in relation to NB-LRR regulation, providing a basis for understanding the functional roles of miRNAs in disease resistance.

This study was conducted to develop the early maturing rice lines with genes conferring resistance to bacterial blight and rice stripe virus to enhance the adaptability in plain area. Unkwang carrying Xa3 was used as a recurrent parent and SR30075 carrying Xa4+xa5+Xa21+Stvb-i was used as a donor parent. RL1(Resistant Line, BC1F7), RL2, RL3, RL4, and RL5(BC2F6) were bred through bio-assay of K3a race inoculation and phenotypic selection of agronomic traits. The presence of introduced genes was confirmed by testing the resistance levels against bacterial blight and rice stripe virus and then double-checked by using DNA marker. RL1 has all target genes, Xa3+xa5+Xa21+Stvb-i. RL2, RL3, and RL5 have Xa3+Xa21+Stvb-i whereas RL4 has only Xa21. Rice lines carrying Stvb-i showed resistance reaction to rice stripe virus. The combinations of bacterial blight resistant genes(Xa3+xa5+Xa21 and Xa3+Xa21) were found to be promising, as the rice lines carrying these genes enhanced a strong resistant reaction against 16 bacterial blight isolates. Also, the inoculation of K3a race did not alter the brown rice yield, ripened grain ratio and kernel quality of brown rice compared to control. Although RL1 containing all the target resistance genes showed excellent resistance performance, it is not suitable to cultivate in plain area due to instability to lodging, 80% yield level than Unkwang, and low grain quality. RL5 backcrossed twice with Unkwang was found to be a promising line due to its effective resistance gene combination, Xa3+Xa21+Stvb-i and good agronomic traits such as stability to lodging, higher yield and quality compared to Unkwang.

자포니카형 조생 찰벼의 벼흰잎마름병 저항성 증진을 위하여 내도복 다수성 조생 찰벼인 상주찰벼 배경에 벼흰잎마름병 저항성 유전자 Xa2, Xa3, xa5, Xa21을 도입한 저항성 계통을 여교배와 벼흰잎마름병 생물검정을 통해 육성하였고, 해당 저항성 유전자를 표지하는 DNA 분자 마커를 이용하여 저항성 유전자 도입 여부를 확인하였으며 우리나라 벼흰잎마름병 균계 네 개와 필리핀 11개 균계를 대상으로 저항성 반응을 조사하였다. Xa2가 도입된 HR24465 계통은 우리나라 K1,K2 균계와 필리핀 race 9a에 저항성을 나타냈으며 Xa3가 도입된 HR24666 계통은 우리나라 K3a와 필리핀 race 6을 제외하고 저항성 및 중도 저항성을 나타냈다. xa5가 도입된 HR24668과 HR24673은 필리핀 race 6을 제외하고 모두 저항성 반응을 나타냈으며 Xa21이 도입된 HR24669는 우리나라 K1 균계와 필리핀 race 10을 제외하고 저항성 및 중도 저항성을 나타냈다. 육성된 계통들은 상주찰벼와 같이 조생이면서 찰벼였고 출수기와 수장, 수수, 정현비율 및 백미수량이 상주찰벼와 차이가 나지 않았다. 상주찰벼 배경에 Xa2, Xa3,xa5, Xa21이 도입된 저항성 계통은 벼흰잎마름병 저항성이 부족한 자포니카형 조생 찰벼의 저항성 강화를 위하여 유용한 육종소재로 활용될 것이다.

MADS-box transcription factor (TF), primarily involved in the floral organ specification with other several aspects of plant growth and development. Whole genome survey of B. rapa revealed 167 MADS-box genes and categorized into MIKCc, MIKC*, Mα, Mβ and Mγ groups based on phylogeny, protein motif structure and exon-intron organizations. MIKCc group belongs 89 genes, which is the highest in number than in any other crops till date. The MIKCc group has further classified into 13 sub-families. In case of chromosomal localization, remarkably 57 MIKCc type MADS-box genes were found in the duplicated segments of B. rapa genome, whereas only 4 M-type genes have resulted from tandem duplications. Besides floral and vegetative tissue expression we also identified MADS-box genes with their male and female gametophyte specific expression in different stages of flower bud development. Furthermore, from a low temperature treated whole genome microarray data set 19 BrMADS genes were found to show variable transcript abundance in two contrasting double haploid lines of B. rapa. Subsequently, the responsive genes were investigated under three abiotic stresses where they showed differential and corresponsive expression patterns. An extensive annotation and transcriptome profiling undertaken in this study might be useful for understanding the involvement of MADS-box genes in stress resistance besides their growth and developmental functions, which ultimately will provide the basis for functional characterization and exploitation of the candidate genes in the genetic engineering study of B. rapa

Numerous environmental stresses, such as abiotic and biotic stresses, cause significant yield loss in crops and can significantly affect their development. Un the field conditions, crops are exposed to a variety of concurrent stresses. Combined high temperature and linked diseases can cause considerable damage that eventually leads to crop death. Hence, this study was conducted to characterize the genes encoding the nucleotide-binding site (NBS) motif obtained from transcriptome profiles of two cabbage genotypes with contrasting responses to heat stress. We selected 80 up-regulated genes form a total of 264 loci, among which 17 were confirmed to be complete and incomplete members of the TIR-NBS-LRR (TNL) class families, and another identified as a NFYA-HAP2 family member. Expression analysis using qRT-PCR revealed that 8 genes showed significant responses to heat shock treatment and F. oxysporum infection. Additionally, in the commercial B. oleracea cultivars with resistance to F. oxysporum, Bol007132, Bol016084, and Bol030522 genes showed dramatically higher expression levels in the F. oxysporum resistant line than the intermediate and susceptible lines. The results of this study may facilitate the identification and development of molecular markers based on multiple stress resistance genes related to heat and fungal stress under field conditions in B. oleracea.

Soybean mosaic virus (SMV), a member of Potyviridae family, is one of the most typical viral diseases and results in yield and quality loss of cultivated soybean. Due to the depletion of genetic resources for resistance breeding, a trial of genetic transformation to improve disease resistance has been performed by introducing SMV-CP and HC-Pro gene by RNA interference (RNAi) method via Agrobacterium-mediated transformation. Transgenic plants were infected with SMV strain G5 and investigated the viral response. As a result, two lines (3 and 4) of SMV-CP(RNAi) transgenic plants and three lines (2, 5 and 6) of HC-Pro(RNAi) transgenic plants showed viral resistance. In genomic Southern blot analysis, most of lines contained at least one T-DNA insertion in both SMV-CP(RNAi) and HC-Pro(RNAi) transgenic plants. Subsequent investigation confirmed that no viral CP and HC-Pro gene expression was detected in two SMV-resistant lines of SMV-CP(RNAi) and three lines of HC-Pro(RNAi) transgenic plants, respectively. On the other hand, non-transgenic plants and other lines showed viral RNA expression. Viral symptoms affected seed morphology, and clean seeds were harvested from SMV-resistant line of SMV-CP(RNAi) and HC-Pro(RNAi) transgenic plants. In addition, strong viral gene expression was detected from seeds of SMV-susceptible non-transgenic plants and SMV-susceptible transgenic lines. When compared the viral resistance between SMV-CP(RNAi) and HC-Pro(RNAi) transgenic plants, soybean transgenic plants with the HC-Pro gene using RNAi strategy showed much stronger and higher frequency of viral resistance.

Rice blast disease caused by the fungal pathogen Magnaporthe oryzae is one of the most destructive rice diseases worldwide. Resistance to rice blast pathogen mostly shows a quantitative trait controlled by several genes. A total of 13 major blast resistance (R) genes were reported in a number of Korean rice varieties using molecular markers. The Pi-ta gene, which locates near to the centromere of chromosome 12, was haplotyping using 1790 accessions including cultivated and wild varieties in previous research. However, the genetic variations of other R genes in rice still not clear. Three R genes, Pi9, Pia, and Pib on chromosome 6, 11 and 2 respectively, were resequenced among 84 accessions of rice core set. Different types of halotype among the 84 accessions were detected. Some new SNPs and InDels found in exon part of R genes were expected to result into amino acid changes following analysis of the genetic code variations, and the germplam in this rice core set which are resistance to blast were explored. We are expecting to develop the new functional markers and incorporate of resistance genes into existing rice cultivars and finally these apply outcomes in breeding rice resistance to blast diseases.

Plants have evolved a set of protecting mechanisms against pathogens, which include secondary metabolites and induced defense responses to pathogen attack. The biological role of purine alkaloids including caffeine is largely unknown. It has been proposed that caffeine confers a resistance against pathogenic bacteria and herbivores. We, in this study, tested direct effects on the growth of rice pathogenic microbes, Xanthomonas oryzae pv. oryzae (Xoo) causing a bacterial leaf blight and Magnaporthe grisea (M. grisea) causing a rice blast. Cell growth of Xoo and M. grisea were significantly retarded in presence of high concentration (2mM) of caffeine. Exogenous caffeine (5mM) induced resistance of wild type rice (cv. Dongjin, susceptible to Xoo and M. grisea) against those pathogens. These results indicated that caffeine enhanced the basal resistance to infection with Xoo. In addition, expression of pathogenesis-related (PR) genes was tested in the caffeine treated rice to elucidate the acquired resistance by caffeine, resulted in induction of PR genes including OsPR1a and OsPrb1. We have generated a transgenic rice producing caffeine by introduction of three N- methyltransferase genes (CaXMT1, CaMXMT1, CaDXMT1) identified from coffee plant. The transgenic rice successfully expressed the three genes, synthesized caffeine up to 5ug/g and showed enhanced resistance to Xoo. We also observed that transcripts of PR genes such as the OsPR1a and OsPrb1 encoding PR-1 type pathogenesis-related protein increased in the caffeine-producing rice. These result showed that caffeine is likely to act a powerful factor to increase level of rice defense as a natural and non-harmful metabolite.

Fungal blast caused Magnaporthe oryzae, bacterial blight caused by Xanthomonas oryzae pv. oryzae (Xoo), bacterial streak caused by Xanthomonas oryzae pv. oryzicola (Xoc) are devastating diseases of rice worldwide. Application of host resistance to these pathogens is the most economical and environment-friendly approach to solve this problem. Some major disease resistance (MR) genes controlling qualitative resistance and quantitative trait loci (QTLs) controlling quantitative resistance are valuable sources for broad-spectrum and durable disease resistance. We have characterized a number of rice MR genes and resistance QTL genes that confer a broad-spectrum or durable resistance to M. oryzae, Xoo, and Xoc. How to efficiently use these genes for rice improvement will be discussed.