Background : Codonopsis lanceolata is a perennial plant of Campanulaceae and mainly distributed in East Asia such as Korea, China, and Japan. C. lanceolata has a unique taste and aroma, and it is rich in minerals such as phosphorus and calcium, and vitamin B1 and B2, so our ancestors used the plant as medicinal herb and edible vegetable. However, systematic cultivation and development of varieties have not been achieved compared to demand or high added value. The genetic diversity and relationship analysis of the plants help to increase the efficiency of breeding through genetic variation. Methods and Results : Ten species of Codonopsis plants were used as materials and DNA was extracted from each 4 individuals per species and quantified at a concentration of 10 ng /㎕. The extracted DNA was pooled by species and PCR was performed using the EST-SSR marker developed based on C. lanceolata in the previous study. PCR amplification was carried out using a denaturation at 94℃ for 30 sec, annealing at 58℃ for 30 sec and extension at 72℃ for 30 sec, repeated for 35 total cycles. The PCR products were separated in a 4% agarose gell at 100 V for 40 min. Conclusion : In this study, C. lanceolata collections was determined among several Codonopsis species using these molecular marker. It is expected that the data of this study can be used as reference for genetic polymorphism analysis and related gene studies of Codonopsis species.

Understanding the genetic variation among landrace collections is important for crop improvement and utilization of valuable genetic resources. The present study was carried out to analyse the genetic diversity and associated population structure of 621 foxtail millet accessions of Korean landraces using 22 EST-SSR markers. A total of 121 alleles were detected from all accessions with an average of 5.5 alleles per microsatellite locus. The average values of gene diversity, polymorphism information content, and expected heterozygosity were 0.518, 0.594, and 0.034, respectively. Following the unweighted neighbor-joining method with arithmetic mean based clustering using binary data of polymorphic markers, the genotypes were grouped into 3 clusters, and population structure analysis also separated into 3 populations. Principal coordinate analysis (PCoA) explained a variation of 13.88% and 10.99% by first and second coordinates, respectively. However, in PCoA analysis, clear population-level clusters could not be found. This pattern of distribution might be the result of gene flow via germplasm exchanges in nearby regions. The results indicate that these Korean landraces of foxtail millet exhibit a moderate level of diversity. This study demonstrated that molecular marker strategies could contribute to a better understanding of the genetic structure in foxtail millet germplasm, and provides potentially useful information for developing conservation and breeding strategies.

The objective of this study was to investigate the expression of bovine luteum expressed sequence tags (ESTs), vascular endothelial growth factor (VEGF), and tumor necrosis factor receptor 1 (TNFR1) and the presence of functional ESTs in the bovine corpus luteum (CL) during different stages of the estrus cycle. Reverse transcription-polymerase chain reaction (RT-PCR) analysis showed a difference in the expression of ESTs during the CL stage. Concentration of ESTs in the CL tissue increased significantly from the mid-luteal stage and decreased thereafter. RT-PCR analysis showed higher levels of the EST genes in the CL of the mid-luteal stage than in other stages, and the same level of expression of VEGF. Immunohistochemistry analysis of the tissue from CL formation to regression showed low cytosol and aggregation of the nucleus. And activity caspase 3 (apoptosis detector) was most strongly detected in the CL1 stage of bovine. During the estrous cycle, the cytosol was magnified and differentiation of the nucleus was clearly manifested. The ESTs affected the CL, and the relationship between VEGF and TNFR1 played a pivotal role for CL development and activation, dependent on the stage of CL. These results suggest local production of ESTs, the presence of functional ESTs in the bovine CL, and that ESTs play a role in regulating the function of cell death in bovine CL.

To successful molecular breeding, identification and functional characterization of breeding related genes and development of molecular breeding techniques using DNA markers are essential. Although the development of a useful marker is difficult in the aspect of time, cost and effort, many markers are being developed to be used in molecular breeding and developed markers have been used in many fields. Single nucleotide polymorphisms (SNPs) markers were widely used for genomic research and breeding, but has hardly been validated for screening functional genes in olive flounder. We identified single nucleotide polymorphisms (SNPs) from expressed sequence tag (EST) database in olive flounder; out of a total 4,327 ESTs, 693 contigs and 514 SNPs were detected in total EST, and these substitutions include 297 transitions and 217 transversions. As a result, 144 SNP markers were developed on the basis of 514 SNP to selection of useful gene region, and then applied to each of eight wild and culture olive flounder (total 16 samples). In our experimental result, only 32 markers had detected polymorphism in sample, also identified 21 transitions and 11 transversions, whereas indel was not detected in polymorphic SNPs. Heterozygosity of wild and cultured olive flounder using the 32 SNP markers is 0.34 and 0.29, respectively. In conclusion, we identified SNP and polymorphism in olive flounder using newly designed marker, it supports that developed markers are suitable for SNP detection and diversity analysis in olive flounder. The outcome of this study can be basic data for researches for immunity gene and characteristic with SNP.

Molecular markers are useful for selecting to include superior character genetic like as strong immune system and rapid growth in fish. The marker is also very important part of breeding technology in Olive flounder (Paralichthys olivaceus). Single nucleotide polymorphisms (SNPs) marker is already in use widely for genomic research and breeding. But this SNPs marker hardly has been validated for screening functional genes in Olive flounder. We study identify single nucleotide polymorphisms (SNPs) on Expressed sequence tag (EST) database, develop usable SNP marker and apply to wild sample and cultured of olive flounder. As a result, Out of total 4.327 ESTs, 693contigs and 514 SNP from total contigs were detected while these substitutions include 297 transitions and 217 transversions. 144 developed markers were applied in 16 samples (wild 8, culture 8), Out of total marker, only 32 markers had detected polymorphic in sample. Polymorphism of 32 markers was observed in the variety genes region involved in immunity and protein synthesis. And the 32 marker were identified 21 transitions, 11 transversions, and indel was not detected in polymorphic SNPs. The analysis on heterozygosity by sample showed 0.34 in wild sample and 0.29 in cultured sample. In conclusion, we was identified SNP and Polymorphism by designed new marker, it supports that development marker is suitable for SNP detection and diversity analysis in Olive flounder. The outcome of this study can be basic data for researches for immunity gene and characteristic with SNP.

Foxtail millet (Setaria italica L.) is the second most widely cultivated species of millet, especially in East Asia and is a tractable experimental model crop for studying functional genomics of millets. However, insufficient researches had been conducted about the foxtail millet germplasm and is significantly impeding its genetic improvement. We attempted to develop EST-derived-SSR (eSSR) markers and utilize them in genetic comparison of germplasm and transferability. A total of 66,027 foxtail millet EST sequences and 42,754 genomic sequence were deduced transcriptom. Approximately 42,000 single tone contigs were generated using DNAstar 5.0 software for redundancy minimization. Nearly 33% of the 14,012 unigenes contained SSRs, but primers were designed for a total of 314 microsatellites concentrating with more than 24 bp of repeats. A total of 314 primers were successfully designed with more than 24 bp of repeats. From these microsatellites, 56 primer pairs were showed polymorphism with over than 15 bp differences among 96 accessions collected from different countries. Polymorphic information content (PIC) value ranged from 0.020 to 0.700 with an average of 0.381 indicating moderate level of informativeness within these EST-SSRs markers. The EST-SSR markers developed in this study will serve as a useful source for genetic studies, such as genetic variability, transferability, association mapping, and molecular breeding

Toxic (high phorbol esters) and nontoxic (low phorbol esters) jatropha accessions cannot be distinguished morphologically. Their seeds must be chemically analyzed through a complex and costly process using HPLC method. EST-SSR markers can be used to classify jatropha accessions with high and low phorbol esters. In this study, ninety-seven EST-SSR markers amplified the genomic DNA and showed polymorphism among 5 high phorbol esters accessions and 10 low phorbol esters accessions. These markers can be further exploited for jatropha improvement through marker-assisted breeding.

본 연구는 감 수집종의 분류 및 품종 육종을 위하여 EST-SSR 마커를 개발해 유전적 유연관계를 분석하고, 형태적 유연관계를 비교 분석하여 DNA 마커의 효율성을 극대화한 연구 결과이다. 경북농업기술원 상주감시험장에서 수집한 42품종을 대상으로 6가지의 양적형질(과실크기, 과고, 과경, 과경굵기, 과경길이, 종자크기)과 19가지의 질적형질(횡단면, 종단면, 골의 정도, 얕은 동심원 균열, 옆모양, 정부열과, 세로홈, 꽃받침 끝 주름, 배꼽 홈길이, 꽃받침 쪽의 홈, 꽃받침 크기)을 사용하여 형태적 유연관계를 분석하였다. 유전적 유연관계를 분석하기 위해 수집한 감에서 cDNA library를 만들어 sequence를 분석한 후, PCR을 통해 얻은 polymorphism이 인정되는 25개의 primer set에서 16개의 EST-SSR primer set를 선발하였다. 수집한 감 42품종의 형태적 유연관계와 개발한 14개의 EST-SSR 마커를 이용하여 유전적인 유연관계를 분석한 결과 형태적 유연관계에서는 여러 그룹이 형성되었지만 coefficient가 0.02 이하로 형성되어 형태적 특성을 사용해 분류하기는 어려웠다. 유전적 유연관계는 coefficient 0.77에서 3개 그룹으로 분류되어, 상주수수감과 상주수꽃감, 밀양반시와 밀양고동시, 영동반시와 영동수시는 각각 같은 그룹으로 분류되었다. 형태적 분석과 유전적 분석의 상관관계를 조사한 결과 형태적 분석의 유사도 거리와 유전적 분석의 유사도 거리 간의 값이 -0.03으로 유의성이 매우 낮게 나왔다. 본 실험에서 얻어진 분자마커는 (EST-SSR 마커) 국내 감육종 효율 증진뿐만 아니라 우수형질을 도입하는데 유용하게 이용할 수 있을 것으로 기대된다.

Genome duplication is an abundance phenomenon and in plant kingdom and consequently formed paralogous region. Korean ginseng (Panax ginseng C.A. Meyer) has a possibility of tetraploid by comparing chromosome numbers of relative species. During development of EST-SSR markers in Korean ginseng, most of primer sets have produced multiple bands in gel electrophoresis. In this study, for identifying origin of multiple bands, five EST-SSR markers showing multi-band were selected and two bands around expected size were sequenced. Sequence comparison classified the multiple bands into individual loci. Two bands can be identified by SNP or InDel variation with number of SSR units. Sequencing result represented that paralogous loci with high similarity were existence caused by recent duplication. One clear band were amplified with newly designed locus specific primer picked from SNP variation. SNP and InDel polymorphism between paralgous loci were useful for identifying each locus. This study will provide better understanding of ginseng genome and will be helpful for development of DNA markers.

EST-SSRs were developed in Brassica napus by database mining. We isolated 7,802 EST-SSRs from the B. napus 643,946 ESTs deposited in the NCBI. With the cut-off value of >10 repeats in di-nucleotide repeats and >7 tri-nucleotide repeats, 303 ESTs were suitable for primer designing for PCR amplification. Of the sixteen possible di-nucleotide combinations, only three types of repeats (AC/GT, AG/CT, and AT/TA) were present among the di-nucleotide EST-SSRs. Whereas, 27 tri-nucleotide repeat motifs from the 64 possible combinations were present. The repeat numbers ranged from 10-15 in di-nucleotide repeats and 7-9 in tri-nucleotide repeat motifs, respectively. By checking PCR amplification in 10 Korean rapeseed breeding lines or cultivars, 234 primer pairs showed successful PCR amplification and 142 of the 234 primer pairs revealed polymorphism among the control cultivars or breeding lines. While the repeat length does not related with the SSR polymorphisms, the repeat motif number showed positive relation with the polymorphism generation by showing higher repeat numbers with higher polymorphisms. We are here presenting the PCR amplifiable primer sequences of the 234 SSR primer pairs to aid in the germplasm management and breeding programs of the B. napus in Korea.

In this study, Expressed Sequence Tag-Simple Sequence Repeat (EST-SSR) analyses were used to clarify the genetic polymorphisms among Korean ginseng cultivars and breeding lines and to classify them into distinct genetic groups. Polymorphic and reproducible bands were produced by 14 primers out of total 30 primers used in this study. Fourteen EST-SSR loci generated a total of 123 bands. Amplified PCR products showed the highly reproducible banding patterns at 110~920 bp. The number of amplified bands for each EST-SSR primers ranged from 2 to 19 with a mean of 8.8 bands. P26 and P35 primers showed 13 and 12 banding patterns, respectively. The number of alleles for each EST-SSR locus ranged from 1.67 to 2.00 with a mean of 1.878 alleles. P34 and P60 primers showed the highest and the lowest genetic polymorphism, respectively. Cluster analysis based on genetic similarity estimated by EST-SSR markers classified Korean cultivars and breeding lines into 4 groups. Group included Gopoong and Chunpoong and 9 breeding lines (55%), group included 2 breeding lines (10%), group included 3 breeding lines (15%), group included Gumpoong and 3 breeding lines (20%). Consequently, the EST-SSR marker developed in this study may prove useful for the evaluation of genetic diversity and differentiation of Korean ginseng cultivars and breeding lines.

This study was carried out to develop expressed sequence tag-Simple sequence repeat (EST-SSR) markers of brown plant hopper resistance gene originated from a rice cultivar ‘Cheongcheong’ and sensitive rice cultivar ‘Nakdong’. Total RNA extracted from the leaves of ‘Cheongcheong’ and ‘Nakdong’ were used to synthesize a cDNA library. As a result of analyzing the cDNA library, EST-SSR sites were found and the EST-SSR primer sets were developed. This study enables to provide effective marker assisted selection (MAS) methods on the selection of white-backed planthopper resistance gene originated from a rice cultivar more simply, quickly and precisely. Furthermore, using this marker’s advantage of deriving from cDNA, it is possible to identity the white-backed planthopper resistance gene.

The olive flounder (Paralichthys olivaceus) is one of the most widely cultured flatfish in Korea and Japan. During development, in a process known as metamorphosis, this fish reorients itself to lie on one side, the body flattens, and the eye migrates to the other side of the body. However, few studies have focused on molecule regulation mechanism of eye development in olive flounder. To reveal the molecular mechanism of eye development, we performed the studies on identification of genes expressed in the eye of olive flounder using EST and RT-PCR strategy. A total of 270 ESTs were sequenced, and 178 (65.9%) clones were identified as known genes and 92 (34.1%) as unknown genes. Among the 178 EST clones, 29 (16.3%) clones were representing 9 unique genes identified as homologous to the previously reported olive flounder ESTs, 131 (73.6%) clones representing 107 unique genes were identified as orthologs of known genes from other organisms. We also identified a kind of eye development associated proteins, indicating EST as a powerful method for identifying eye development-related genes of fish as well as identifying novel genes. Further functional studies on these genes will provide more information on molecule regulation mechanism of eye development in olive flounder.

Perilla is a genus as a member of the mint family Lamiaceae which is known to contain lots of volatile metabolite. Perilla has been called as ‘deulkae’ indicating ‘wild sesame’ that means it has been maintained in Korea with long history. It has been very friendly used as edible oil and as fresh leaf vegetable. Perilla oil is valued for its medicinal benefit because it contains best amounts of unsaturated fatty acids, especially for the alpha-linolenic acid, known to omega-3 fatty acid, among all of the plant oils. It also include many beneficial phytochemicals. However, little study is conducted on their genetics. Here, we announce construction of well normalized and full length enriched-perilla cDNA library from a whole plant of one cultivar ‘Youngho-deulkae’ and their sequence characterization to provide useful resources for genetics, breeding and metabolite engineering. By sequencing of 5,760 cDNA clones, we 5,438 high quality EST sequences. Sequence trimming and assembly resulted 3,995 unigenes which consists 1,004 contigs and 2,991 singletones. Unigenes that showed little homology at the DNA sequence level with known genes in other plants even though they showed similarity at the protein domain level based on BLASTN, BLASTX, and TBLASTX. This study may provide good resources for initiation of further genomics, comparative genomics, functional genomics such as metabolic engineering and molecular breeding.