• Korean Journal of Plant Taxonomy
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• v.34 no.1
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• pp.43-61
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• 2004

Molecular taxonomic studies were conducted to evaluate interspecific relationships in Korean Viola 34 taxa including two Japanese populations using RAPD(randornly amplified polymorphic DNA), ISSR(inter simple sequence repeat) and PCR-RFLP(restriction fragment length polymorphism) analysis. Only six and four primers out of 40 arbitrary and 12 ISSR primers were screened for 34 taxa, and were revealed 70 (98.6%) and 28 (96.6%) polymorphic bands, respectively. Fifteen restriction endonucleases produced 80 restriction sites and size variations from the large single copy region of cpDNA, 16 (20%) of which were polymorphic. The separate analyses from the RAPD, ISSR and PCR-RFLP data were incongruent in the relationships among 34 taxa, but combined data was in accordance with previous infrageneric classification system based on morphological characters, especially the subsection and series level. Section Chamaemelanium placed between subsect. Patellares and Vagimtae of section Nomimium was not formed as a distinct group. Viola alb ida complex including three very closely related taxa was recognized independent group within subsect. Patellares in combined data tree. This result strongly suggested that they should be treated to series Pinmtae. RAPD analysis was very useful to clarify the interspecific relationships among the species of Korean Viola than ISSH and PCR-RFLP analyses.

• Journal of Plant Biotechnology
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• v.47 no.4
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• pp.298-308
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• 2020

Genetic diversity among Thaumatococcus daniellii populations in the southwestern region of Nigeria were assessed using morphometric and molecular markers to determine the population structure and existing genetic relationship for its improvement, conservation and sustainable utilisation. Populations from five locations in each of the six states were used for the study. Morphometric data were collected on folia characters and analysed for variability. Genome DNA was isolated from the plant leaf and amplified by polymerase chain reaction with inter-simple sequence repeat markers (ISSR) to determine the allelic polymorphism, marker effectiveness and genetic relationship of the population. The results showed significant variations in petiole length and leaf dimensions of the populations within and across the states. These morphometric traits are the major parameters that delimit the populations and they correlated significantly at P≤0.05. Analysis of the electrophoregram showed that the ISSR markers are effective for the diversity study. A total of 136 loci were amplified with an average of 7.16 loci per marker, 63.2% of the loci were polymorphic. The Principal Coordinate Analysis revealed that seven factors accounted for 81.6% of the variation and the dendrogram separated the populations into two major groups at a genetic distance of 10 (about 90% similarity) with sub-groups and clusters. Most populations within the state had a high degree of similarity, nonetheless, strong genetic relationship exists among populations from different states. The close relationship between populations across the states suggests a common progenitor, which are likely separated by ecological or geographical isolation mechanisms.

• KOREAN JOURNAL OF CROP SCIENCE
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• v.49 no.1
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• pp.69-72
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• 2004

Somaclonal variation, defined as phenotypic and genetic variations among regenerated plants from a parental plant, could be caused by changes in chromosome structure, single gene mutation, cytoplasm genetic mutation, insertion of transposable elements, and DNA methylation during plant regeneration. The objective of this study was to evaluate DNA variations among somaclonal variants from the cotyledonary node culture in soybean. A total of 61 soybean somaclones including seven $\textrm{R}_1$ lines and seven $\textrm{R}_2$ lines from Iksannamulkong as well as 27 $\textrm{R}_1$ lines and 20 $\textrm{R}_2$ lines from Jinju 1 were regenerated by organogenesis from the soybean cotyledonary node culture system. Field evaluation revealed no phenotypic difference in major agronomic traits between somaclonal variants and their wild types. AFLP and SSR analyses were performed to detect variations at the DNA level among somaclonal variants of two varieties. Based on AFLP analysis using 36 primer sets, 17 of 892 bands were polymorphic between Iksannamulkong and its somaclonal variants and 11 of 887 bands were polymorphic between Jinju 1 and its somaclonal variants, indicating the presence of DNA sequence change during plant regeneration. Using 36 SSR markers, two polymorphic SSR markers were detected between Iksannamulkong and its somaclonal variants. Sequence comparison amplified with the primers flanking Satt545 showed four additional stretches of ATT repeat in the variant. This suggests that variation at the DNA level between somaclonal variants and their wild types could provide basis for inducing mutation via plant regeneration and broadening crop genetic diversity.

• International Journal of Industrial Entomology and Biomaterials
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• v.19 no.2
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• pp.249-253
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• 2009

Raily ecorace of Indian tasar silkworm is wild in nature and distributed abundantly in dense deciduous forest on Shorea robusta (Sal) in Bastar ($17^{\circ}4'$ and $20^{\circ}34'$ N, $80^{\circ}15'$ and $82^{\circ}15'$ E and altitude ranging from 150 to 1200 mMSL) forest ranges of Chhattisgarh, India. It is represented by about 20 populations. Out of those, eleven populations showed intra- as well as inter- population variability based on phenotypic expression and also in major economic traits viz. cocoon weight, shell weight, filament length and denier. Genetic diversity in these eleven populations was studied using Inter-Simple Sequence Repeat (ISSR) markers. The band profiles generated with eight ISSR primers have depicted variation in band size. All the primers exhibited polymorphism which is an indicative of the genetic variation in individual Raily silkworm. Among the populations, total polymorphism recorded was 76%. The population genetic aspects assessed through POPGENE software package are discussed in the paper. Nei's gene diversity (h) ranged from 0.194 to 0.337 exhibiting high heterozygosity. Relevance of the present study is of high significance in formulating conservation strategies and sustainable utilization of the economically important Raily ecorace of Antheraea mylitta.

• Korean Journal of Plant Resources
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• v.28 no.6
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• pp.697-703
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• 2015

Twenty apple germplasm accessions from the Korean Genebank were successfully cryopreserved using two-step freezing to back up genetic resources maintained by field collections. This study examined the morphological and genetic stability of cryopreserved dormant apple buds that were stored in liquid nitrogen, and then rewarmed and regrown. Whole plants were regenerated directly from dormant buds through budding without an intermediary callus phase. The cryopreserved buds produced high levels of shoot formation (76.2-100%), similar to those of noncryopreserved buds (91.3-100%), with no observed differences between cryopreserved and noncryopreserved materials. Three of the twenty cryopreserved apple germplasm accessions were used to assess morphological and genetic stability. No differences in morphological characteristics including shoot length, leaf shape, leaf width/length ratio, and root length were observed between controls (fresh control and noncryopreserved) and cryopreserved plantlets. The genetic stability of regenerants (before and after cryopreservation) was investigated using inter simple sequence repeat (ISSR) markers. The ISSR markers produced 253 bands using four primers, ISSR 810, SSR 835, ISSR 864, and ISSR 899. These markers showed monomorphic banding patterns and revealed no polymorphism between the mother plant and regenerants before and after cryopreservation, suggesting that cryopreservation using two-step freezing does not affect the genetic stability of apple germplasm. These results show that two-step freezing cryopreservation is a practical method for long-term storage of apple germplasms.

• Molecules and Cells
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• v.25 no.2
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• pp.205-210
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• 2008

To develop molecular markers linked to the $L^4$ locus conferring resistance to tobamovirus pathotypes in pepper plants, we performed AFLP with 512 primer combinations for susceptible (S pool) and resistant (R pool) DNA bulks against pathotype 1.2 of pepper mild mottle virus. Each bulk was made by pooling the DNA of five homozygous individuals from a T10 population, which was a near-isogenic $BC_4F_2$ generation for the $L^4$ locus. A total of 19 primer pairs produced scorable bands in the R pool. Further screening with these primer pairs was done on DNA bulks from T102, a $BC_{10}F_2$ derived from T10 by back crossing. Three AFLP markers were finally selected and designated L4-a, L4-b and L4-c. L4-a and L4-c each underwent one recombination event, whereas no recombination for L4-b was seen in 20 individuals of each DNA bulk. Linkage analysis of these markers in 112 $F_2$ T102 individuals showed that they were each within 2.5 cM of the $L^4$ locus. L4-b was successfully converted into a simple 340-bp SCAR marker, designated L4SC340, which mapped 1.8 cM from the $L^4$ locus in T102 and 0.9 cM in another $BC_{10}F_2$ population, T101. We believe that this newly characterized marker will improve selection of tobamovirus resistance in pepper plants by reducing breeding cost and time.

• Journal of Life Science
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• v.21 no.3
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• pp.468-472
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• 2011

DNA marker is a powerful tool for plant genetics and breeding. In this study, 995 SSR markers were employed with chilling resistant cucumber, known as 'NC76', and chilling susceptible cucumber, known as 'GY14'. Using 2% agarose gel electrophoresis, 145 SSR markers were identified as length variation markers between 'NC76' and 'GY14'. The SSR markers that showed no length polymorphism were then screened using high resolution melting analysis technique and additional 30 polymorphic SSR markers were identified. As a preliminary evaluation for mapping, 20 markers among these 175 markers were employed to a $F_2$ population of 'NC76' x 'GY14' cross. Linkage analysis revealed 13 markers that joined into six linkage groups and seven markers that remained unlinked. This result indicates that these 175 markers could be used for construction of a genetic map using a cross between 'NC76' and 'GY14' for further investigation in developing markers related to resistance to chilling in cucumbers.

• Korean Journal of Breeding Science
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• v.42 no.5
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• pp.502-506
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• 2010

Soybean seed possesses about 15 allergenic proteins recognized by IgEs from soy-sensitive human. The allergenic impact of soybean proteins limit its extensive usage in a broad range of processed foods. Soybean protein P34 or Gly m Bd 30k of the cysteine protease family is one of the major allergen of the soybean seed. P34-null soybean, PI567476, was identified among soybean (Glycine max & Glycine soja Sieb. and Zucc) of approximately 16,226 accessions from USDA soybean germplasm screened. Also, for P34 gene (Williams 82; whole genome sequence cultivar) and P34 null gene (PI567476) comparative analysis of sequences listed in the NCBI database showed the presence of a SSLP (Simple Sequence Length Polymorphism) of 4 base pair. So, a SSLP marker was designed to reveal the polymorphism of the locus. In this study, a population of 339 $F_2$ recombinant inbred lines generated by cross between Taekwang (Glycine max) and PI567476 was used to select $F_{2:3}$ plant of a P34 null gene. The result separation rate Taekwang type, heterozygous type and PI567476 type were shown in 85: 187: 67 since single gene is concerned in as the separation rate of 1:2:1 in $X^2{_{0.05}}=5.99$, df=2. In future, selected plant will identify protein level, whether P34 null protein is equal to P34 null gene.

• Journal of Microbiology and Biotechnology
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• v.18 no.4
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• pp.624-630
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• 2008

Terminal restriction fragment length polymorphism (T-RFLP) is a method that has been frequently used to survey the microbial diversity of environmental samples and to monitor changes in microbial communities. T-RFLP is a highly sensitive and reproducible procedure that combines a PCR with a labeled primer, restriction digestion of the amplified DNA, and separation of the terminal restriction fragment (T-RF). The reliable identification of T-RF requires the information of nucleotide sequences as well as the size of T-RF. However, it is difficult to obtain the information of nucleotide sequences because the T-RFs are fragmented and lack a priming site of 3'-end for efficient cloning and sequence analysis. Here, we improved on the T-RFLP method in order to analyze the nucleotide sequences of the distinct T-RFs. The first method is to selectively amplify the portion of T-RF ligated with specific oligonucleotide adapters. In the second method, the termini of T-RFs were tailed with deoxynucleotides using terminal deoxynucleotidyl transferase (TdT) and amplified by a second round of PCR. The major T-RFs generated from reference strains and from T-RFLP profiles of activated sludge samples were efficiently isolated and identified by using two modified T-RFLP methods. These methods are less time consuming and labor-intensive when compared with other methods. The T-RFLP method using TdT has the advantages of being a simple process and having no limit of restriction enzymes. Our results suggest that these methods could be useful tools for the taxonomic interpretation of T-RFs.

• International Journal of Industrial Entomology and Biomaterials
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• v.30 no.1
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• pp.6-16
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• 2015

This study established the genetic characterisation of 10 microsporidian isolates infecting non-mulberry silkworms (Antheraea assamensis and Samia cynthia ricini) collected from biogeographical forest locations in the State of Assam, India, using PCR-based markers assays: inter simple sequence repeat (ISSR) and random amplified polymorphic DNA (RAPD). A Nosema type species (NIK-1s_mys) was used as control for comparison. The shape of mature microsporidian spores were observed oval to elongated, measuring 3.80 to $4.90{\mu}m$ in length and 2.60 to $3.05{\mu}m$ in width. Fourteen ISSR primers generated reproducible profiles and yielded 178 fragments, of which 175 were polymorphic (98%), while 16 RAPD primers generated reproducible profiles with 198 amplified fragments displaying 95% of polymorphism. Estimation of genetic distance coefficients based on dice coefficients method and clustering with un-weighted pair group method using arithmetic average (UPGMA) analysis was done to unravel the genetic diversity of microsporidians infecting Indian muga and eri silkworm. The similarity coefficients varied from 0.385 to 0.941 in ISSR and 0.083 to 0.938 in RAPD data. UPGMA analysis generated dendrograms with two microsporidian groups, which appear to be different from each other. Based on Euclidean distance matrix method, 2-dimensional distribution also revealed considerable variability among different identified microsporidians. Clustering of these microsporidian isolates was in accordance with their host and biogeographic origin. Both techniques represent a useful and efficient tool for taxonomical grouping as well as for phylogenetic classification of different microsporidians in general and genotyping of these pathogens in particular.