• Journal of Forest and Environmental Science
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• v.25 no.2
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• pp.93-100
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• 2009

In Malaysia, Labisia pumila Benth & Hook f, popularly known as 'Kacip Fatimah' has been used traditionally to treat various elements of the woman's health in Malay community. The objective of this study was to develop randomly amplified polymorphic DNA (RAPD) based DNA markers for the identification of L. pumila and to distinguish its three varieties from each other. Total DNA from nine accessions of L. pumila was extracted by CTAB method and polymerase chain reactions (PCR) were carried out to amplify the segments of DNA using different primers to develop DNA barcode using RAPD technique. To find out variety-specific DNA marker/s, twenty different 10-mer primer sequences with annealing temperature from 36-$40^{\circ}C$ were evaluated in triplicate. Out of 20 random primers, two primers (OPA-1 and OPA-2/A10) were selected which produced reliable RAPD band patterns. To have DNA based handle, two RAPD amplification products were cloned and sequenced to determine the identity of the DNA. RAPD analysis using two random primers generated 72 discrete bands ranging in size 200 bp-3,000 bp. Fifty nine of these were polymorphic loci (82%) and thirteen were non-polymorphic loci (18%). A total of 32 bands polymorphic loci (72%) were amplified with primer OPA-1 and analyzed by cluster analysis and UPGMA (Unweighted Pair Group Method with Arithmetic) to present a dendogram depicting the degree of genetic relationship among nine accessions of L. pumila. Our results shows the reasonable genetic diversity among the L. pumila varieties and within varieties; and two RAPD marker sequences obtained could be used to identify L. pumila at species level.

• Asian-Australasian Journal of Animal Sciences
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• v.19 no.9
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• pp.1234-1239
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• 2006

Murrah and NiliRavi are the important North Indian buffalo breeds occupying the prominent position of being the highest milk producers. These breeds are more or less similar at morphological as well as physiological levels. The technique of RAPD-PCR was applied in the present study to identify a battery of suitable random primers to detect genetic polymorphism, elucidation of the genetic structure and rapid assessment of the differences in the genetic composition of these two breeds. A total of 50 random primers were screened in 24 animals each of Murrah and NiliRavi buffaloes to generate RAPD patterns. Of these, 26 (52%) primers amplified the buffalo genome generating 263 reproducible bands. The number of polymorphic bands for the 26 chosen RAPD primers varied from 3 (OPG 06 and B4) to 26 (OPJ 04) with an average of 10.1 bands per primer and size range of 0.2 to 3.2 kb. DNA was also pooled and analyzed to search for population specific markers. Two breed specific RAPD alleles were observed in each of Murrah (OPA02 and OPG16) and NiliRavi (OPG09) DNA pools. RAPD profiles revealed that 11 (4.2%) bands were common to all the 48 individuals of Murrah and NiliRavi buffaloes. Pair-wise band sharing calculated among the individual animals indicated considerable homogeneity of individuals within the breeds. Within breed, band sharing values were relatively greater than those of interbreed values. The low genetic distance (Nei's) value (0.109) estimated in this study is in accordance with the origin and geographical distribution of these breeds. The RAPD analysis indicated high level of genetic similarity between these two important North Indian buffalo breeds.

• Restorative Dentistry and Endodontics
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• v.29 no.5
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• pp.430-438
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• 2004

This study evaluated the influence of application time of self-etching primers on microtensile bond strength (${\mu}$TBS) to dentin using three self-etching primer adhesive systems. Dentin surfaces were exposed from forty-eight human molars. They were conditioned with three self-etching primers (Clearfil SE Bond [SE], Unifil Bond [UF], Tyrian SPE + One Step Plus [TY]) and different primining times (10s, 20s, 30s and 40s). Composite resins were bonded to dentin surfaces and specimens were made. ${\mu}TBS$ was tested and statistically compared using by one-way ANOVA and Tukey's Test. The results of this study presented that priming time for 10s in SE and UF groups and for 30s and 40s in TY group was highly decreased ${\mu}TBS$ to dentin.

• Korean Journal of Medicinal Crop Science
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• v.13 no.3
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• pp.109-113
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• 2005

To analyze the genetic relationship 8 accessions of Ostericum koreanum Kitakawa, random amplified polymorphic DNA(RAPD) analysis was performed using 60 Operon primers. The 25 primers out of 60 random primers were amplified DNA by PCR using genomic DNA of O. koreanum. Eighty-five (49.1%) among 173 bands derived from 25 primers showed poly morphism, On the basis of similarity coefficient analysis by UPGMA, 8 accessions of O. koreanum Kitakawa could be classified into three groups at the similarity coefficient value of 0.71. Group I contained three accessions (Nam Gangwhal), Group II contained one accession (Nam Gangwhal) and Group III contained four accessions (Buk Gangwhal), The range of total genetic similarity coefficient value of 8 accessions of O. koreanum Kitakawa was $0.63{\sim}0.96$. Buk Gangwhal was flowered 18 to 26 days earlier than Nam Gangwhal, and Nam Gangwhal leaf stalk was thin and long as bolting rate high compared with Buk Gangwhal.

• Journal of the Korean Society of Environmental Restoration Technology
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• v.7 no.5
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• pp.94-99
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• 2004

This study was carried out to find out what is the optimum conditions for RAPD of Zelkova serata. We changes the factors what affect to PCR band patterns, as a result, we established the optimum conditions as follows; template DNA 100mg, Primer 0.25uM, dNTP 100mM, Taq polymerase 1.0u, and total reaction volume was filled up to 10uL with distilled water. As the amount of primers went higher, PCR reaction rates were lowered. This reason was cause by exhaustion of primers during initial reaction. The amount of dNTP didn't showed noticable differtations between the range, but the optimum amount was 100mM for efficiency. Taq polymerase 1.0 unit was the best in the range. As the concentration of polymerase were increased, many non-specific bands were appeared, In primer selection, most Openron Random Primers are amplified in this experiment. The primers GC contents were 60, and set A, B, C, D, E, X were tested. Thermal cycler(ASTEC PC808, Japan) condition was, $95^{\circ}C$, 5min, initial denaturation, $94^{\circ}C$, 20sec, denaturation, $37^{\circ}C$, 40sec, annealing, $72^{\circ}C$, 1min, extention, 45cycle repeated and final extention $72^{\circ}C$10min.

• The Plant Pathology Journal
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• v.20 no.4
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• pp.302-307
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• 2004

Three pairs of specific primers were developed for rapid and precise RT-PCR detection of three seed-transmissible viruses, namely Peanut clump virus (PCV, Pecluvirus), White clover mosaic virus (WCIMV, Potexvirus) and Carrot red leaf virus (CaRLV, Luteovirus). Each primer set was found in conserved region through multiple sequence alignment in the DNAMAN. Total nucleic acids extracted from PCV-, WCMV-, and CaRLV-infected seeds and healthy plants were used for RT-PCR detection using each virus-specific primer, Sizes of PCV, WCIMV, and CaRLV PCR products were 617bp (PCV-uni5 and PCV-uni3 primers), 561bp (WCMV-CP5 and WCMV-CP3 primers), and 626bp (CL1-UP and CL2-DN primers); which corresponded to the target sizes. Nucleotides sequences of each amplified cDNA were confirmed which belonged to the original virus. This study suggests that these virus-specific primer sets can specifically amplify viral sequences in infected seeds. Thus, they can be used for specific detection of three viruses (PCV, WCMV and CaRLV) from imported seed samples for plant quarantine service.

• Journal of Ginseng Research
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• v.17 no.2
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• pp.153-158
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• 1993

The study was carried out for comparison of variants and development of genetic markers using Randomly Amplified Polymorphic D사A (RAPD) analysis method. The ginseng variants used were as follows: Chungkyung-Chong, Hwangskoog-Chong, KG101 selected by the pureline selection method, and 6 kinds of Jakyung-Chong strains Uinjakyung, Jakyung-Chong 81783, Jakyung-Chong 847913, Jaky tong-Chong 79742, Jinjakyung of USSR, and Mimaki of Japan). Four of 10 RAPD primers showed the distinctive polymorphism among 9 ginseng variants and lines, and were selected for more detailed polymorphic analysis. The sequences of 4 selected primers were TGCCGAGCTG (Primer#2), AATCGGGCTG (#4), GAAACGGGTG (U7), and GTGACGTAGG (#8). All primers produced several common bands among the strains. However, when primer # 2 was applied, the electrophoregram showed the specific band at 1.8 kb region in Chungkyung-Chong, Hwangskoog- chong, and KG101, and 1 kb in the Jakyung-Chong 847913. In primer #4, 1.1 kb band was shown in Chungkyung-Chong, Hwangskoog-Chong, KG101, and Jakyung-Chong 79742. In primer # 7, 700 bp band was appeared in Jakyung-Chong 81783 and Jinjakyung of USSR In primer # 8, 800 bp band was observed only in Mimaki, comparing to another strains. When Similarity Index (SI) was calculated, Chungkyung-Chong and Hwngskoog-Chong, and Jakyung- chong 81783 and Jinjakyung of USSR showed the most close SI, 0.11 and 0.08, respectively. The data of KG101, which showed the SI of 0.13 with the group of Chungkyung-Chong and Hwangskoog-Chong, coincided with the fact that it was released from Hwangskoog-Chong by breeding process. The data of Jakyung strains indicated the significant variation among the strains. From these results, RAPD analysis method could be succesively applied to the classification and genetic analysis for breeding of Korean ginseng.

• Korean Journal of Plant Resources
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• v.23 no.5
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• pp.458-464
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• 2010

Twelve garlic cultivars collected from Korea and Mongolia were evaluated genetic similarity and diversity by RAPD method using oligo-nucleotide random primers. Genomic DNA isolated from twelve garlic cultivars were amplified by polymerase chain reaction using 143 primers, and 55 primers showed polymorphic DNA bands. Among a total of 187 bands amplified by 55 primers, 128 polymorphic bands were subjected to analysis for genetic relationship of garlic cultivars. Garlic cultivars were classified into three groups, such as group-I corresponded to Euiseong, Seosan, Samchuk and Yecheon-A, Yechun-B, Euiseong-norang, Jeongsun, Namdo, Yookback and Danyang cultivars, and group-II to Mongolia and group-III to Daeseo cultivars. Thirty DNA bands showing unique specificity to the specific cultivars are likely to be useful for identification of garlic local cultivars as DNA markers.

• Korean journal of applied entomology
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• v.37 no.1
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• pp.23-30
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• 1998

Restriction fragment length polymorphism (RFLP) of a DNA has been a useful tool for analyzing genetic variation. This research was performed to establish an RFLP analytic method on the mitochondrial DNA (mtDNA) of the beet armyworm, Spodoptera exigua (Hiibner). To do this, total size of the mtDNA was measured and polymerase chain reaction (PCR) primers were selected. Its mitochondrial genome size was ca. 16kb. From a serial PCR test of 29 primers refered to the compilation of Simon et al. (1994), 22 primers were selected to amplify its mtDNA fragments. These primers resulted in short (300-700 bp) or long (1000-2000 bp) DNA products which represented a total or partial sequence of each of CO-I, CO-11, Cyt-B, ND-1, 12s rRNA, 16s rRNA, and some tRNAs. PCR-RFLP was performed in some variable mtDNA regions with 8 kinds of 4bp recognizing restriction enzymes. Different populations from Andong, Kyungsan, and Sunchun did not show any restriction site polymorphisms but had some length variation in certain regions of mtDNA.

• Journal of Life Science
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• v.9 no.6
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• pp.671-676
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• 1999

RAPD analysis using random primers were tried to evaluate the genetic variation and diversity of the nine garlic cultivars including two foreign varieties. Thirty-two primers out of 70 primers screened were used to amplify genomic DNA of garlic cultivars using polymerase chain reaction(PCR). Among a total of 151 bands amplified by 32 primers, 125 polymorphic bands were subjected to analysis for genetic relationship of garlic cultivars. The estimated size of amplified PCR products were in the range of 932 to 4,060 base pairs. Nine garlic cultivars were classified into two groups, such as group I corresponded to Changnyung and Hungary cultivars, and group II, Namdo, Sandong from China, Yecheon, Euiseong, Youngweol, Danyang, Jeongsun cultivars, with the genetic distance value of 0.271. The major ecological types of garlics, so called southern and northern types, was grouped in the genetic distance value of 0.200. The results presented in this study suggest that RAPD analysis are likely to be useful for identification of cultivars and evaluation of genetic origin in garlics.