• The Plant Pathology Journal
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• 제34권3호
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• pp.199-207
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• 2018

Tomato chlorosis virus (ToCV) is a whitefly-transmitted and phloem-limited crinivirus. In 2013, severe interveinal chlorosis and bronzing on tomato leaves, known symptoms of ToCV infection, were observed in greenhouses in Korea. To identify ToCV infection in symptomatic tomato plants, RT-PCR with ToCV-specific primers was performed on leaf samples collected from 11 tomato cultivating areas where ToCV-like symptoms were observed in 2013 and 2014. About half of samples (45.18%) were confirmed as ToCV-infected, and the complete genome of 10 different isolates were characterized. This is the first report of ToCV occurring in Korea. The phylogenetic relationship and genetic variation among ToCV isolates from Korea and other countries were also analysed. When RNA1 and RNA2 are analysed separately, ToCV isolates were clustered into three groups in phylogenetic trees, and ToCV Korean isolates were confirmed to belong to two groups, which were geographically separated. These results suggested that Korean ToCV isolates originated from two independent origins. However, the RNA1 and RNA2 sequences of the Yeonggwang isolate were confirmed to belong to different groups, which indicated that ToCV RNA1 and RNA2 originated from two different origins and were reassorted in Yeonggwang, which is the intermediate point of two geographically separated groups.

• 농업과학연구
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• 제43권2호
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• pp.221-231
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• 2016

Microsatellites, which are sequences of repetitive short nucleotides, are abundant in the genome and have relatively many alleles at a locus. Hence, microsatellite markers are used in various research areas such as medicine, agriculture, and biology. Thanks to recent advanced techniques and databases associated with microsatellite marker development, foreign research relying on microsatellite markers is increasing in various study areas. In this study, by analyzing microsatellites-related articles published during 2000-2014 from eight Korean national journals representing zoology, botany, genetics, ecology and environmental science, breeding science, and forest science ('Animal Cells and Systems', 'Journal of Plant Biology', 'Genes and Genomics', 'Korean Society of Environment and Ecology', 'Korean Journal of Breeding Science', 'Journal of Agricultural Science, Chungnam National University', 'Journal of Korean Forest Society' and 'Forest Science and Technology'), we found that the number of articles and diversity of study subjects and objects have increased considerably. However, there are fewer applications of microsatellites in the national forest science area. During 2000-2014 in 'Journal of Korean Forest Society', the percentage of articles dealing with microsatellite markers was found to be the lowest with 4.2% among articles focusing on PCR-based markers including RAPD, AFLP, and ISSR. However, in 'Canadian Journal of Forest Research' and 'Forest Ecology and Management', microsatellite marker articles were represented at their highest with 69.2% and 76.2%, respectively. Given the advantages of microsatellite markers, the publication of research papers using microsatellites should be increased in Korean forest science journals to the level of studies published in prominent international journals.

• 한국작물학회:학술대회논문집
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• 한국작물학회 2017년도 9th Asian Crop Science Association conference
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• pp.89-89
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• 2017

A gene encoding squalene synthase from grain amaranth was cloned and characterized. The full-length cDNA was 1805-bp long and contained a 1248-bp open reading frame encoding a protein of 416 amino acids with a molecular mass of 47.6 kDa. Southern blot analysis revealed that the A. cruentus genome contained a single copy of the gene. Comparison of the cDNA and genomic sequences indicated that the amaranth SQS gene had 12 introns and 13 exons. All of the exons contributed to the coding sequence. The predicted amino acid sequence of the SQS cDNA shared high homology with those of SQSs from several other plants. It contained conserved six domains that are believed to represent crucial regions of the active site. We conducted qRT-PCR analyses to examine the expression pattern of the SQS gene in seeds at different developmental stages and in several tissues. The amaranth SQS gene was low levels of SQS transcripts at the initial stage of seed development, but the levels increased rapidly at the mid-late developmental stages before declining at the late developmental stage. These findings showed that the amaranth SQS is a late-expressed gene that is rapidly expressed at the mid-late stage of seed development. In addition, we observed that the SQS mRNA levels in stems and roots increased rapidly during the four- to six-leaf stage of development. Therefore, our results showed that the expression levels of SQS in stem and root tissues are significantly higher than those in leaf tissues. In present study provides useful information about the molecular characterization of the SQS clone isolated from grain amaranth. Finally, a basic understanding of these characteristics will contribute to further studies on the amaranth SQS.

• 원예과학기술지
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• 제33권4호
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• pp.550-558
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• 2015

To analyze the linkage relationships among molecular markers recently reported to be linked to onion (Allium cepa L.) Ms, a restorer-of-fertility locus, in onion (Allium cepa L.), three single nucleotide polymorphism markers were converted into cleaved amplified polymorphic sequence (CAPS) markers based on onion transcriptome sequences and the rice genome database. Analysis of the recombinants selected from 4,273 segregating plants using CAPS and other linked markers demonstrated the jnurf13 and jnurf610 markers to perfectly co-segregate with the Ms locus. In contrast to jnurf13, the jnurf610 marker was not in perfect linkage disequilibrium with the Ms locus in diverse breeding lines. Thus, the jnurf13 marker and the marker for identification of cytoplasm types were utilized to enhance the efficiency of onion breeding through four applications. First, 89 maintainer lines containing the normal cytoplasm and homozygous recessive Ms genotypes were successfully identified from 100 breeding lines. Second, these two molecular markers were used to analyze the main sources of male-fertile contaminants frequently found in the male-sterile parental lines during F1 hybrid seed production. The majority of the contaminants contained heterozygous Ms genotypes, indicating that pollen grains harboring the dominant Ms genotype may have been introduced during propagation of the maintainer lines. Therefore, the genetic purity of the two maintainer lines was analyzed in the third application, and the results showed that both maintainer lines contained 13-21% off-types. Finally, the two markers were used to increase the seed yield potentials of two open-pollinated varieties containing sterile cytoplasms by removing the plants harboring homozygous recessive and heterozygous Ms genotypes.

• 식물조직배양학회지
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• 제22권3호
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• pp.149-155
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• 1995

국내에서 분리 동정한 오이모자이크 바이러스(CMV)로부터 외피단백질(CP) 유전자를 분리하였고 이의 염기서열을 결정한 결과 129번째 아미노산이 proline으로서 mosaic symptom을 보이고 염기 및 아미노산 서열의 유사성을 비교한 결과 subgroup I (type I)에 속한다는 것을 확인하였다. 분리한 CP 유전자를 식물형질전환용 운반체에 삽입하여 담배 재배종인 NC82와 바이러스 증식용인 Samsun에 잎절편을 통하여 형질전환시켰다. 형질전환된 담배의 DNA를 분리하여 Southern blotting과 PCR을 수행한 결과 대부분의 형질전환체에 CP 유전자가 삽입되였음을 확인하였고 T$_1$ 종자를 kanamycin이 첨가된 배지에서 후대 항생제 저항성 검정을 실시한 결과 1개의 CP 유전자가 삽입된 경우가 50%에 달했고 2개와 3개의 유전자가 삽입된 경우도 각각 39%와 11%에 달했다.

• Journal of Plant Biotechnology
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• 제45권1호
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• pp.45-54
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• 2018

This research was conducted to study the gene expression of coffee (Coffea arabica L.) seedlings under salt stress condition. A solution of five percent ($2.3dS\;m^{-1}$) deep sea water was used for the salt treatment, and it was thereby compared to normal irrigation water ($0.2dS\;m^{-1}$) used for the control treatment. The mRNA was extracted from the leaves of the coffee seedlings for a comprehensive analysis. In this study, a total of 19,581 genes were identified and aligned to the reference sequences available in the coffee genome database. The gene ontology analysis was performed to estimate the number of genes associated with the identified biological processes, cellular components and molecular functions. Among the 19,581 genes, 7369 (37.64%) were associated with biological processes, 5909 (30.18%) with cellular components, and 5325 (27.19%) with molecular functions. The remaining 978 (4.99%) genes were therefore grouped as unclassified. A differential gene expression analysis was performed using the DESeq2 package to identify the genes that were differentially expressed between the treatments based on fold changes and p-values. Namely, a total of 611 differentially expressed genes were identified (treatment/control) in that case. Among these, 336 genes were up-regulated while 275 of the genes were down-regulated. Of the differentially expressed genes, 60 genes showed statistically significant (p < 0.05) expression, 44 of which were up-regulated and 16 which were down-regulated. We also identified 11 differentially expressed transcription factor genes, 6 of which were up-regulated and rest 5 genes were down-regulated. The data generated from this study will help in the continued interest and understanding of the responses of coffee seedlings genes associated with salinity stress, in particular. This study will also provide important resources for further functional genomics studies.

• Asian-Australasian Journal of Animal Sciences
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• 제18권5호
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• pp.723-727
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• 2005

Ghrelin is a novel growth-hormone-releasing acylated peptide, which has been purified and identified in rat stomach. In the present study, the full-length sequence of bovine ghrelin cDNA was cloned by RT-PCR. The bovine ghrelin cDNA sequence derived in the present study included a 348 bp open reading frame and a 137 bp 3'UTR. The putative amino acid sequence of bovine prepro-ghrelin consisted of 116 amino acids, which contained the 27-amino acid ghrelin. The sequence analysis of the bovine ghrelin gene revealed that an intron existed between Gln$^{13}$ and Arg$^{14}$ of ghrelin. This exon-intron boundary matched the GT-AG rule of the splicing mechanism. Compared with rats, which have two tandem CAG sequences in the 3'end of intron, bovine ghrelin genome has only one CAG sequence. Therefore, although rats can produce 28 amino acid-ghrelin and 27 amino acid-des-Gln$^{14}$-ghrelin by alternative splicing, ruminant species, including bovines, might be able to produce only one type of ghrelin peptide, des-Gln$^{14}$-ghrelin. The influence of aging on plasma ghrelin concentration was also examined. Plasma ghrelin concentration increased after birth to approximately 600 days of age, and then remained constant.

• BMB Reports
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• 제34권4호
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• pp.322-328
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• 2001

Excision and amplification of pre-determined, large genomic segments (taken directly from the genome of a natural host, which provides an alternative to conventional cloning in foreign vectors and hosts) was explored in human cells. In this approach, we devised a procedure for excising a large segment of human genomic DNA, the iNOS gene, by using the Cre/loxP system of bacteriophage P1 and amplifying the excised circles with the EBNA-1/oriP system of the Epstein-Barr virus. Two loxP sequences, each of which serves as a recognition site for recombinase Cre, were integrated unidirectionally into the 5'-UTR and 3'-UTR regions of the iNOS gene, together with an oriP sequence for conditional replication. The traps-acting genes cre and EBNA-1, which were under the control of a tetracycline responsive $P_{hcmv^*-1}$ promoter, were also inserted into the 5'-UTR and 3'-UTR regions of the iNOS gene, respectively, by homologous recombination. The strain carrying the inserted elements was stably maintained until the excision and amplification functions were triggered by the induction of cre and EBNA-1. Upon induction by doxycycline, Cre excised the iNOS gene that was flanked by two ZoxP sites and circularized it. The circularized iNOS gene was then amplified by the EBNA-1/oriP-system. With this procedure, approximately a 45.8-kb iNOS genomic fragment of human chromosome 17 was excised and successfully amplified in human cells. Our procedure can be used effectively for the sequencing of unclonable genes, the functional analysis of unknown genes, and gene therapy.

• Journal of Microbiology and Biotechnology
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• 제23권12호
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• pp.1708-1716
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• 2013

Conventional molecular detection methods cannot distinguish between viable and dead Escherichia coli O157 cells. In this study, the loop-mediated isothermal amplification (LAMP) method combined with propidium monoazide (PMA) treatment was developed to selectively detect viable E. coli O157 cells. Four primers, including outer primers and inner primers, were specially designed for the recognition of six distinct sequences on the serogroups (O157) of the specific rfbE gene of the E. coli O157 genome. PMA selectively penetrated through the compromised cell membranes and intercalated into DNA. Amplification of DNA from dead cells was completely inhibited by $3.0{\mu}g/ml$ PMA, whereas the DNA derived from viable cells was amplified remarkably within 1 h by PMA-LAMP. Exhibiting high sensitivity and specificity, PMA-LAMP is a suitable method for evaluating the inactivation efficacy of slightly acidic electrolyzed water in broth. PMA-LAMP can selectively detect viable E. coli O157 cells. This study offers a novel molecular detection method to distinguish between viable and dead E. coli O157 cells.

• Journal of Crop Science and Biotechnology
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• 제10권3호
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• pp.175-184
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• 2007

Two subspecies, japonica and indica, have been reported in rice, which differ in several ecotypic traits. However, reproductive barriers in hybrid progenies between subspecies have been major obstacles in breeding programs using inter-subspecific hybridization. As the first step to elucidate the reproductive barriers, we developed subspecies-specific(SS) STS markers in this study. A total of 765 STS primers were designed through comparing DNA sequences at every $2{\sim}3$cM interval between japonica and indica rices, which are available at Web DBs such as IRGSP, NCBI, TIGR, and GRAMENE, and tested for subspecies-specificity using 15 indica and 15 japonica varieties of diverse origin. Of them, 67 STS markers were identified as SS STS markers and their subspecies-specificity scores were estimated. The SS markers were dispersed throughout the genome along chromosomes. Of them, 64 SS markers were mapped on an RIL population derived from a Dasanbyeo(indica)/TR22183(japonica) cross. Genomic inclination of RILs was evaluated based on the genotyping with different types of markers. Association test between markers and segregation distortion revealed that segregation distortion might not be the cause of generating SS markers. The SS markers will be applicable to estimate the genomic inclination of varieties or lines and to study the differentiation of indica and japonica, and ultimately to breed true hybrid rice varieties in which desirable characters from both subspecies are recombined.