Plant breeding requires genetic diversity of useful traits for crop improvement. EMS-induced mutation is practiced to generate mutations at loci regulating economically important traits and/or to knock out the genes to elucidate their functions. The present study was aimed to induce mutations in a Korean local land race Capsicum annuum ‘Yuwol-cho’. This accession is pungent and also has advantage to mature early. A total of about 1,500 M2 families were screened and three non-pungent mutants were identified and crossed with wild type ‘Yuwol-cho’. After phenotyping of F2 population for pungency, MutMap approach will be used to identify the genes controlling the pungency in mutants. In addition to this, another C. annuum accession “Micro-Pep” was used to develop a mutant population. Micro-Pep is a small, pungent pepper generally used as ornamental purpose. Having compact growth habit, and small size, it has advantage to handle and utilize easily in mutation study and molecular research. On the basis of preliminary experiment 1.3% of mutagen was used for treatment of pepper seeds and 30% less germination percentage was observed in EMS treated seeds in comparison to control seeds. A total of 4,674 M1 plants are grown under greenhouse condition and M2 population will be studied for characterization of phenotypic variation including fruit color and pungency. Newly constructed mutant populations will be valuable assets for identification of functional genes and molecular breeding of pepper.

Arsenic (As) is accumulated in rice grain due to environmental reasons such as polluted ground water and soil, and As toxicity constitutes a serious threat to human health. However, the accurate information required for understanding As-responsive mechanisms remain mostly unknown in rice. Here, we performed the comparative genome-wide transcriptome analysis between As tolerance type (ATT) rice mutant induced by γ-irradiation and its wild type (WT). As compared to WT after As treatment of 150 ppm, ATT exhibited the phenotypic differences such as vigorous growth in shoots and root hairs, and low accumulation of H2O2 in rice roots. In transcriptome analysis, we found between WT and ATT that As toxicity commonly affected to inhibit gene regulations involved in photosynthesis, mitochondrial electron transport and lipid biosynthesis metabolism. While, many genes associated with cysteine synthesis metabolism considerably up regulated in both As-treated plants. Additionally, we found the potential As tolerance-related genes involved in abiotic stress-responsive mechanism and RNA-protein synthesis for protein degradation and modification. To further analyzes the genetic variations of As-responsive genes, the DNA polymorphic DEGs associated with oxidoreductase significantly distributed in ATT more than in WT.

Goami 2 is now well known as its extinguishing endosperm characteristics - it is far from the wild type, Ilpum, a premium taste Korean japonica cultivar. The endosperm of Goami 2 is high in fat, protein, and indigestible carbohydrate contents. One of the most extraordinary endosperm characteristic of Goami 2 is high level of amylose content, even though the hulled rice (brown rice) is totally opaque. There have been many studies to address the unique physico-chemical properties and possible usages as a healthy and functional food ingredient, especially, the high-amylose rice had a positive effect on lowering the blood glucose response in obesity and type 2 diabetes. Genetic analysis by using 44 SSR markers, crude linkage map (3~5 anchor markers per chromosome) was then constructed based on the genotypes detected among 112 F2 progenies derived from Goami 2 / Milyang 23 showed that major chromosomal regions on Chromosome 2 responsible for the variation of amylose contents. M2-53 on Chromosome 2 explains the highest variation and this region has not been reported as a putative QTL for amylose contents yet. More closely markers for application to breeding program can be developed using MutMap or Re-sequence methods.

To understand the molecular mechanism of leaf morphogenesis in rice, ethylmethane sulfonate (EMS) treated Ilpoom mutant line with semi-narrow and adaxially rolled leaf phenotype was identified. The leaf rolling character is said to be more advantageous under high temperature and heat stress, and play as one of the defensive mechanisms. The F1 plants, generated from a cross of Ilpoom and mutant, showed normal phenotype. Genetic analysis of its F2 population suggested that the mutation was controlled by a single recessive gene with segregation ratio of 3:1. Using F2 mapping population derived from a cross of Ilpoom mutant and Milyang23, each chromosomes were screened with STS markers by the bulked segregant analysis (BSA) method. The candidate region was detected to a long arm of chromosome 1 near the centromeric region. Fine mapping of the locus is currently conducted. Moreover, other morphological characterizations of the mutant plants were identified. Cytological analysis of the leaf suggested that deformation of the bulliform cells led to the smaller size and less number of the bulliform cells, and caused leaf rolling trait.

Carotenoid isomerase (CRTISO) catalyzes the isomerization of prolycopene to all-trans-lycopene in the carotenoid biosynthetic pathway. We isolated two full-length cDNA gene, CuCRTISO and CuCRTISO-like, from Citrus unshiu. To confirm whether these two genes have the function of the carotenoid isomerase, The full-length cDNA of CuCRTISO and CuCRTISO-like gene, respectively, were fused with 35S promoter and NOS terminal region and then transformed into tomato CRTISO mutant, Tangerine, which shows orange fruit due to lack of carotenoid isomerase activity. The mature fruit color of the transgenic line expressing CuCRTISO gene changed from orange to red, which was similar to the fruit color of the tomato “Money Maker”. We also carried out HPLC analysis to detect all-trans lycopene, which is produced from prolycopene by carotenoid isomerase. In the transgenic line expressing CuCRTISO the all trans lycopene was detected from mature fruit but in the tangerine mutant several prolycopenes were detected from it. On the other hand, the transgenic line expression of CuCRTISO gene retained the orange-color fruit at the mature stage as Tangerine mutant. These studies indicate that the CuCRTISO gene has a function of carotenoid isomerase and also plays a role of it in other plant species, and that the CuCRTISO-like gene might be not enough to produce the all trans lycopene or has a another unknown function(s).

Panax ginseng C.A Meyer is commonly used in Asian traditional medicine to treat a variety of diseases. Ginsenosides are glycosylated triterpenes, referred to saponins, have been especially noted as active compounds contributing to the various efficacy of ginseng. In this study, we are trying to select high saponin content of ginseng lines from the gamma irradiated adventitious roots. Recently, we have generated several mutant ginseng lines improving ginseniside content by gamma radiation. The mutant lines were selected by phenotypes and ginsenoside content (HPLC analysis) of the irradiated adventitious root lines. However, the ginsenoside content of the mutant lines was not sufficient for commercial use and the selection method was not suitable for large scale of mutant line selection. In this study, we are testing Fourier transfer infrared spectroscopy (FT-IR) as a new selection method of mutant lines in Panax ginseng. About 5,000 pieces of Panax ginseng adventitious roots were exposed to gamma radiation (60Co). Irradiation dosages were 0, 25, 50 and 70Gy. Survival rate of the irradiated samples was evaluated by counting the number of survival main roots after 5 weeks culture in the solid MS medium with NAA, IAA and 5% sucrose. In present, we are collecting the survived adventitious root lines (about 900 lines) from the gamma irradiated ginseng roots for FT-IR and HPLC analysis. After analysis of FT-IR and HPLC, we will assess the suitability of the FT-IR as a screening method for the preparation of mutant lines in ginseng.

To identify novel signaling components involved in regulation of plant responses to phosphate (Pi) starvation, we screened an Arabidopsis T-DNA activation tagging library for mutants with altered Pi-starvation responses. Here, we report the identification and characterization of novel activation-tagged mutant involved in Pi starvation signaling in Arabidopsis. The hpd (hypersensitive to Pi deficiency) mutant exhibits enhanced phosphate uptake and altered root architectural change under Pi starvation compared to wild type. Expression analysis of auxin-responsive DR5::GUS reporter gene in hpd mutant indicated that both auxin biosynthesis and auxin translocation under Pi starvation are suppressed in hpd mutant plants. Impaired auxin translocation in roots of hpd mutant was attributable to abnormal root architecture changes in Pi starvation conditions. Mis-regulation of auxin translocation in hpd mutant was further confirmed by analysis of expression patterns of auxin efflux carrier proteins, PIN-FORMED (PIN) 1, 2, and 3 fused with GFP. Not only expression levels but also expression domains of PIN proteins were altered in hpd mutant in response to Pi starvation. Molecular genetic analysis of hpd mutant revealed that the mutant phenotype is caused by the lesion in ENHANCED SILENCING PHENOTYPE4 (ESP4) gene whose function is proposed in mRNA 3’-end processing. The results propose that mRNA processing plays crucial roles in Pi homeostasis as well as developmental reprograming in response to Pi deprivation in Arabidopsis.

In the rice inflorescence development, timing of inflorescent meristem abortion, conversion of the rachis branch meristem to the terminal spikelet meristem and shift to lateral meristem identity determine the overall architecture of the rice panicle (keda-Kawakatsu et al. 2009). Cheng et al. (2011) reported that quantitative trait loci (QTLs) have major effects on panicle apical abortion in rice. However, there have been very few reports about panicle tip mutants. Therefore, this research is conducted to fine map mutant gene and perform functional analysis of mutant gen. Hwacheongbyeo (japonica rice) seed was treated with ethyl methane sulfonate (EMS) for inducing mutation. Two F2 population (Japanica mutant crossed with wild type and Japanica mutant crossed with Milyang 23, Indica type) were established for Phenotyping and genomic analysis. STS markers in crop molecular Breeding laboratory. Additional STS markers for fine mapping were developed based on the Nipponbare genome sequence (http://rgp.dna.affrc.go.jp/blast/runblast.html). All F2 generations showed the segregation of normal plants and mutant following a ratio of 3:1 suggesting the mutant phenotype is caused by a single recessive gene. Initial BSA test made using STS markers confirmed the mutant gene is found in the long arm of chromosome 8. Panicle tip mutant gene, pnt has pleotropic effect which has been manifested in significant reduction of tiller development starting from late stage of vegetative growth and pronounced effect on possession of stay green nature of the rice during the vegetative stage of development. The only significant difference observed within panicle traits is the number of spikelet on primary branch and spikelet fertility. The first primary branch which contain aborted spikelet and elongated distance between spikelet is the most affected structure in the panicle.

This study was carried out to determine the amount of lignin, cellulose, and hemicellulose in six kenaf cultivars during different harvesting stages. Three mutant cultivars (Jangdae, Jeokbong and Baekma), two original cultivars (Jinju, C14), and one Chinese cultivar (Auxu) were planted on May 14, 2013. Four harvesting times were made at intervals of 20 days from 15 July to 16 September, 2013. The overall growth characters of mutant cultivar ‘Jeokbong’ such as plant height, stem diameter, flowering time, and dry mass were similar with those of the original variety. The mutant cultivar ‘Baekma’ occurred 10-day late flowering in comparison with the original variety and also displayed higher dry mass than the original variety. Jinju, Auxu and Jangdae, mid-late maturing kenaf cultivars, had high dry weight compared to early maturing cultivars such as Jeokbong, Baekma and C14. In all cultivars, the lignin contents were increased by a late harvest. The Mid-late maturing kenaf cultivars showed high lignin content in comparison with those of the early maturity cultivars. There were no significant differences of cellulose, and hemicellulose content between the cultivars, however cellulose content in stems of these kenaf cultivars were significantly decreased by a late harvest. These results may provide valuable information to assist the parental selection of kenaf breeding.

Kenaf (Hibiscus cannabinus L.) native to Africa can be used as fiber, food, feedstock and bio plastic. This study was carried out to evaluate the mineral, amino acid and vitamin contents of six selected kenaf cultivars which are enable to produce seed under Korean circumstance. The leaves of three mutant cultivars (Jangdae, Jeokbong and Baekma), two original cultivars (Jinju, C14) and one Chinese cultivar (Auxu) were harvested at flowering time. Mineral components of kenaf leaves, such as calcium, potassium, and mineral, did not showed significant differences among the cultivars. As major amino acids including proline and phenylalanine, significant differences were found in these kenaf cultivars. The Auxu cultivar contained the highest amount of essential amino acid (Phenylalanine, Leucine, Isoleucine, Valine, Methionine and Lysine). The amount of vitamin displayed significant differences such as vitamin E and vitamin K among these cultivars. Especially, Jangdae cultivar contained the highest amount of vitamin E and vitamin K. Thus, these data suggested that Jangdae and Auxu is the most desirable cultivar containing high amount of vitamin and amino acid.

Salt and drought stresses affect virtually every aspect of plant physiology and metabolism and thus limiting the productivity of crop plants worldwide. Salt and drought tolerance and adaptation in rice has been improved by engineering various genes related to transcription, signaling, accumulation of antioxidants and compatible solutes etc. Previously, we have produced 2,000 non-GM mutants induced by Tos17 in rice. We analyzed >2,000 flanking sequences of newly transposed Tos17 copies by the adaptor-ligation PCR method. We also identified significantly up- or down-regulated genes under drought, salt, or ABA stress in rice based on expression microarray data, which previously were performed from leaf at different developmental stages and conditions. For screening and characterizing the salt or drought tolerance mutations by extensive phenotypic analysis as well as the functional analysis of genes, we selected 133 mutant lines. To evaluate rice phenotypic traits under abiotic stress condition, we plan to investigate phenomics, which integrates technologies such as photonics, biology, computers, and robotics.

Leaves are the organ for photosynthesis, respiration and transpiration, and have a major effect on crop yield. Therefore, leaf shape and structure are important agronomic traits in breeding for ideal type plant. We obtained a new abaxially rolled leaf mutant from Ilpum(Oryza sativa ssp. japonica) by the treatment of ethyl methane sulfonate(EMS). The abaxially rolled leaf mutant showed reduced plant height and panicle length, increased tiller number and panicle number than Ilpum. LRI(Leaf rolling index) analysis showed that the mutant have high value compared to the wild-type. In cross section analysis, the mutant was observed to have increased of bulliform cell number and size, and led to the outcurved leaf rolling. The phenotypes of the F1 plants derived from the cross between the mutant and Ilpum were normal. In F2 population, segregation ratio between the wild type and the mutant was 3:1. This genetic analysis indicated that leaf rolling is controlled by single recessive gene. Bulked segregant analysis(BSA) and genetic mapping were conducted using F2 population derived from the cross between mutant and Milyang23(Oryza sativa ssp. indica). According to the results, the gene was located on the long arm of chromosome2. Fine mapping is in progress.

작물학적 특성을 신속히 보완하며 확충할 수 있는 이점을 가지고 있는 돌연변이 육종을 통해 육성된 유용형질발현 돌연변이계통들의 다양한 작물학적 특성과 병해충 저항성은 유용한 유전자원으로 평가되고 있으며, 다양한 작물학적 특성 및 병해충 저항성에 대한 돌연변이 계통의 유용형질들을 교배를 통해 고품질 벼의 보완을 위해 이전하는 것이 용이한 이점을 가지고 있다. 농촌진흥청 국립식량과학원에서는 국내 육성 자포니카 벼품종인 남일에 아지드화나트륨을 돌연변이원으로 활용하여 단간이며 도열병 저항성 돌연변이 후대계통인 Namil(SA)- bl5를 육성하였다. 본 연구는 염색체상에서 Namil(SA)-bl5의 단간 특성과 도열병 저항성에 관련된 유전좌위를 탐색하고자 수행하였다. 주요 연구 결과로 Namil(SA)-bl5와 밀양23호간에서 유래한 F2 94 개체로부터 단간에 대한 표현형을 조사하고 68개 SSR 마커의 유전자형을 검정하여 연관분석(association analysis)을 수행한 결과 목표 유전좌위는 6번 염색체 중하단 부위와 7번 염색체 하단 부위로 간장의 단축은 주동유전자적 조절에 의한 것으로 추정되었다. 또한, 분자마커의 유전자형과 도열병저항성 간의 연관성평가를 수행한 결과를 보면, Namil(SA)- bl5의 도열병 저항성에 관여하는 주동 유전좌위는 12번 염색체 중단의 RM1337 좌위에 위치하는 것으로 추정되었다. 추후 목표 유전좌위에 대한 마커밀도를 선택적으로 높여 고밀도 유전자지도 작성함으로 초정밀 분자표지인자를 개발할 계획이다.

작물학적 특성을 신속히 보완하며 확충할 수 있는 이점을 가지고 있는 돌연변이 육종을 통해 육성된 유용형질발현 돌연변이계통들의 다양한 작물학적 특성과 병해충 저항성은 유용한 유전자원으로 평가되고 있으며, 병해충 저항성에 대한 돌연변이 계통의 유용형질들을 교배를 통해 고품질 벼의 보완을 위해 이전하는 것이 용이하다. 농촌진흥청 국립식량과학원에서는 국내 육성 자포니카 벼 품종인 남일에 EMS를 돌연변이원으로 활용하여 도열병 및 벼멸구 저항성 돌연변이 후대계통인 Namil(EMS)-bl10,bph1를 육성하였다. 염색체 상에 Namil(EMS)-bl10,bph1의 도열병 및 벼멸구 저항성에 관련된 유전자좌위를 표지하기 위해 Namil(EMS)-bl10,bph1 x 밀양23호로부터 유래한 F2 97 개체로부터 도열병과 벼멸구에 대한 저항성을 검정하고 60개 SSR 마커의 유전자형을 검정하여 연관성분석(association analysis)을 수행하였다. 분자마커의 유전자형과 저항성 간의 연관성분석 결과, Namil(EMS)- bl10,bph1 의 도열병 및 벼멸구 저항성에 관여하는 주동유전자위는 12번 염색체 중단의 RM1337 및 RM0277에 각각 표지 되었다.

We have generated 383 independent transgenic lines for genetically modified (GM) rice that contained GPD, UtrCSP, BrTSR15 and BrTSR53 genes overexpression constructs under the control of the constitutive CaMV 35S promoter. TaqMan copy number assay was determined inserted T-DNA copy number. Also FSTs analysis was isolated from 203 single copy T-DNA lines of transgenic plants and sequence mapped to the rice chromosomes. In analyzing single copy lines, we identified 95 FSTs, among which 37 (38.9%) were integrated into genic regions and 58 (61.1%) into intergenic regions. About 27 homozygous lines were obtained through multi-generations of planting, resistance screening and TaqMan copy number assay. To investigate the transgene expression patterns, quantitative real-time PCR analysis was performed using total RNAs from leaf tissue of single copy, intergenic region of T-DNA insertion and homozygous T2 plants. The mRNA expression levels of the examined transgenic rice were significantly increased in all of the transgenic plants. In addition, myc-tagged 35S::BrTSR15 and 35S::BrTSR53 transgenic plants were displayed higher levels of transgene protein than WT plants. These results may be useful for producing of large-scale transgenic plants or T-DNA inserted mutants in rice.

Plastochron and phyllotaxy are important traits to determine plant architecture. A plastochron 1-6 mutant was derived from japonica rice cultivar Koshihikari treated with ethylmethane sulfonate (EMS). The plastochron 1-6 mutant showed reduced plant height, shorter internode length, smaller and more leaves than Koshihikari, its wild type. In addition, the mutant has abnormal panicle, abnormal seed and upper node tillers. The F1 plants between mutant and Koshihikari were normal. In F2 population, segregation ratio between the wild type and mutant was fitted to 3:1. This genetic analysis indicated that plastochron 1-6 is controlled by a single recessive gene. Bulked segregant analysis revealed that the gene was located on chromosome 10. Through sequencing analysis, we found that plastochron 1-6 mutant had a single nucleotide transition occurred in the first exon of LOC_Os10g26340 (encoding P450 CYP78A11). This work was supported by a grant from the Next-Generation BioGreen 21 Program (Plant Molecular Breeding Center No. PJ008125), Rural Development Administration, Republic of Korea.

such as small plant size, small genome, short life cycle, self-fertility, and etc. Moreover, Brachypodium standard line Bd21 had already been sequenced and the data of which now have been available to the public. The development of next-generation sequencing technologies has allowed the discovery of large numbers of genome-wide DNA polymorphisms. Brachypodium standard line Bd21 was exposed to chronic gamma radiation. M2 1376-1 line was less stained with phloroglucinol compared to wild-type, which indicated reduced lingin content. Also, it’s filial generations showed dwarf phenotype. Genomic DNA was extracted from the M3 plant and was used to construct a whole-genome re-sequencing library for using Illumina HiSeq2500. The trimmed reads were aligned to the Brachypodium reference genome sequence (http:// www.phytozome.net). SICKLE, BWA, and Picard were used for accurate variant detection. More than 110 M reads were generated and 96.53% of them were mapped. This represents ~35 fold coverage. As a result, mutant specific SNVs, Insert/Deletions, and non-synonymous mutations were obtained. Moreover, non-synonymous mutations were identified from 5 lignin biosynthesis genes (Bradi1g31320, Bradi3g52350, Bradi5g21550, Bradi3g22980, Bradi5g14720). The obtained results will be incorporated in development of biofuel crops.

A sugary mutant with low total starch and high sugar content was compared with its wild type Sindongjin for grain-filling caryopses. In the present study, developing seeds of Sindongjin and sugary mutant from the 11th day after flowering (DAF) were subjected to RNA sequencing (RNA-Seq). A total of 30,385 and 32,243 genes were identified in Sindongjin and sugary mutant. Transcriptomic changes analysis showed that 7,713 differentially expressed genes (DEGs) (log2 Fold change ≥1, false discovery rate (FDR) ≤ 0.001) were identified based on our RNA-Seq data, with 7,239 genes up-regulated and 474 down-regulated in the sugary mutant. A large number of DEGs were found related to metabolic, biosynthesis of secondary metabolites, plant-pathogen interaction, plant hormone signal transduction and starch/sugar metabolism. Detailed pathway dissection and quantitative real time PCR (qRT-PCR) demonstrated that most genes involved in sucrose to starch synthesis are up-regulated, whereas the expression of the ADP-glucose pyrophosphorylase small subunit (OsAGPS2b) catalyzing the first committed step of starch biosynthesis was specifically inhibited during the grain-filling stage in sugary mutant. Further analysis suggested that the OsAGPS2b is a considerable candidate gene responsible for phenotype of sugary mutant.

In this study, the concentrations of isoflavones, anthocyanins and total phenol content (TPC) in 19 soybean mutant lines changed seed coat color from yellow to black or brown were determined. Among 19 soybean mutant lines, 5 soybean mutant lines with black pigment were detected 3 anthocyanins (delphinidin-3-O-β-D-glucoside, D3G; cyaniding 3-O-β -D-glucoside, C3G; petunidin 3-O-β-D-glucoside, Pt3G). The highest concentration of anthocyanins among 5 soybean mutant lines was D-16 (1280.0 ± 19.4 mg/100g seed coat) derived from cv. Danbaek. Although isoflavone contents of all soybean mutant lines showed lower levels compared to original cultivars, glycitein was detected only 5 soybean mutant lines (DP-37-2, DP-38, DP-39, DP-40, and DP-41 derived from cv. Daepung). In TPC of 19 soybean mutant lines, DP-10 was increase levels compared to original cultivar, while DP-37-2, DP-40, and DP-41 were decrease levels of TPC. Using reduction of DPPH, we measured the free radical scavenging activity (FRSA) among 19 soybean mutant lines. Five black and 4 brown soybean mutants showed significant increase in FRSA. On the basis of these results, it was concluded that gamma irradiation may change the isoflavone, anthocyanin, and total phenol contents of soybean. These mutant lines using in this study can be useful for the breeding of soybean varieties altering the nutritional values.

일품벼 유래 돌연변이 고아밀로스 품종인 고아미2호, 고아 미3호, 고아미4호의 저항전분 함량과 전분 특성을 분석한 결 과, 저항전분 함량은 11.87～13.69%로 일품(0.36%)에 비해 상당히 높은 유의적인 함량 차이를 나타냈다. 총식이섬유 함 량 역시 고아밀로스 품종이 일품벼 대비 2～4배 정도로 높았 다. X-선 회절도에 의한 아밀로펙틴 결정 구조 차이에서 일품 의 회절각도(2θ)는 15.0, 17～18, 22～23에서 강한 피크를 보여 전형적인 A형 양상을 보인 반면, 고아미2호, 고아미3호, 고아미4호는 B형의 형태를 나타냈다. 고아밀로스 품종은 아 밀로펙틴 짧은 사슬(A, DP 6-12) 비율이 24.5～26.2%로 일 품(40.3%)에 비해 유의적으로 낮은 분포 비율을 나타냈다. 저항전분 함량이 많은 고아미2호, 고아미3호, 고아미4호는 신 속점도계에 의한 호화 점도 특성에서 호화개시온도가 높고 강하점도는 유의적으로 낮은 특성을 보였다. 저항전분과 쌀의 이화학성분 특성과의 상관성 분석 결과, 쌀의 아밀로스 함량 과 식이섬유 함량이 많을수록, 호화개시온도가 높을수록, 아밀로펙틴의 장쇄사슬이 많을수록 저항전분의 함량이 높아지 는 정의 상관성을 나타냈다.