• Asian Pacific Journal of Cancer Prevention
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• v.16 no.5
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• pp.2073-2080
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• 2015

Background: The human papillomavirus (HPV) and its variants show wide geographical distribution and have been reported to cause cervical lesions. With cervical neoplasia as the leading cancer in Indian women, the aim of the present study was to evaluate the multiple infection HPV type distribution and variant genotypes in cervical samples from the coastal Karnataka region, India. Materials and Methods: A total of 212 samples were screened by nested polymerase chain reaction using PGMY9/11 and GP5+/6+ primers. HPV positive samples were sequenced to identify the types and a phylogenetic tree was constructed using the neighbor-joining method. Results: Sequence analysis identified a total of 14 HPV types distributed in 20%, 73.3% and 82.5% of non-malignant, pre-malignant [low grade squamous intraepithelial lesion (LSIL) and high grade squamous intraepithelial lesion (HSIL)] and cervical cancer samples. The distribution of high risk HPV in cancer samples was HPV 16, 76.4%, HPV18, 11.7%, HPV81, 2.9%, HPV31, 1.4%, HPV35, 1.4% and HPV 45, 1.4%. Multiple infections were observed in 11.8% of tumor samples with HPV 16 contributing to 62.5% of cases. In non-malignant samples, 20% of HPV positive samples were detected with HPV16, 82.3%, HPV33, 5.8% and HPV58, 5.8% and very low incidence of multiple infections. Comparative phylogenetic analysis of HPV variants identified 9 HPV sequences as new papillomavirus species, predominantly classified as European lineage type. Conclusions: The findings for HPV infections associated with progression of cervical cancer in coastal Karnataka region and HPV variant analysis provide baseline data for prevention and HPV vaccination programs.

• International Journal of Industrial Entomology and Biomaterials
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• v.18 no.1
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• pp.18-27
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• 2009

B. thuringiensis strain LY-99 belonging to subsp. alesti (H3a3c), was isolated from Chinese tobacco warehouse and showed significantly high toxicity to Plutella xylostella. For the identification of the cry1-type genes from B. thuringiensis LY-99, an extended multiplex PCRrestriction fragment length polymorphism (PCRRFLP) method was established by using two pairs of universal primers based on the conserved regions of the cry1-type genes to amplify around 2.4 kb cry1-type gene fragments. Then the DNA fragment was cloned into pGEM-T Easy vector and digested with EcoRI and EcoRV enzymes. Through this method, a known cry1-type gene was successfully identified from the reference strain, B. thuringiensis subsp. alesti. In addition, the RFLP patterns revealed that B. thuringiensis LY-99 included a novel cry1A-type gene in addition to cry1Aa, cry1Ac, cry1Be and cry1Ea genes. The novel cry1A-type gene was designated cry1Ah2 (Genbank accession No DQ269474). An inverse PCR method was used to amplify the flank regions of cry1Ah2 gene. Finally, 3143 bp HindIII fragment from B. thuringiensis LY-99 plasmid DNA including 5' region and partial ORF was amplified, and sequence analysis revealed that cry1Ah2 gene from LY-99 showed 89.31% of maximum sequence similarity with cry1Ac1 crystal protein gene. In addition, the deduced amino acid sequence of Cry1Ah2 protein shared 87.80% of maximum identity with that of Cry1Ac2. This protein therefore belongs to a new class of B. thuringiensis crystal proteins.

• Reproductive and Developmental Biology
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• v.29 no.1
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• pp.9-13
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• 2005

This study was performed to analyze the sequence variation in mtDNA D-loop and their effects on milk and milk fat production in Holstein cows. The analyzed sequences were compared with previously published sequences from other cattle breeds (GenBank J01394). PCR was performed to amplify a total of 964 bp between nucleotide 15758 and 383 within D-loop region of mtDNA using specific primers. Thirty five polymorphic sites by nucleotide substitution were found in mtDNA. The frequencies of positions at 106, 169, 16057, 16231 and 16255 nt with high levels of sequence polymorphism were 0.090, 0.555, 0.055, 0.090 and 0.050, respectively. The substitution effect at 169 nt was found significant on milk production, and substitution effect at 16118, 16139 and 16302 nt was highly significant (p<0.1) on milk fat production. Polymorphism of mtDNA sequence in D-loop region might be useful for the analysis of cytoplasmic genetic variation and associations with the other economically important traits and maternal lineage analysis in Holstein cows.

• Research in Plant Disease
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• v.9 no.3
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• pp.107-115
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• 2003

Chinese yam (Dioscorea opposita cv. Jang-Ma) plants showing necrotic mosaic symptom were collected from their growing fields in Andong, Euisong, Gunwi and Daegu, Korea. Direct negative stainning method by electron microscope showed filamentous particles of about 660 nm in length. Immunosorbent electron microscopy (ISEM) revealed filamentous particles of 660nm decorated with antiserum of Chinese yam necrotic mosaic virus (ChYNMV). The virues purified partially were used to isolate viral RNA as template for RT-PCR to amplify about 1.2 kbp of 3'-terminal region (coat protein, 3'-UTR) with ChYNMV specific and oligo-dT primers. Amino acids sequences of amplified CP genes revealed that the viruses shared 97.9% similarity with ChYNMV (AB044386) wh ich was known as the member of Macluravirus. So the viruses from Chinese yam (D. opposita cv. Jang-Ma) plants were identified as ChYNMV. Comparing the CP amion acid sequences of ChYNMV strains with other macluraviruses such as Cardamon mosaic virus (CdMV), Narcissus latent virus (NLV) and Maclura mosaic virus (MacMV) revealed that N-terminal was the most varialbe region and conserved regions were present within the genus Macluravirus.

• Korean Journal of Medicinal Crop Science
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• v.8 no.2
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• pp.102-108
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• 2000

Morphological characteristics of nine Epimedium spp was analyzed on the basis of six morphological characters. To analyse the genetic relationship among Epimedium spp., polymerase chain reaction (PCR) was performed with total genomic DNA of 17 Epimedium spp. by using random 8 primers. The genetic diversity and genetic distance among nine Korean collections, seven Japanese and one China collection were used to generate a dendrogram showing phylogenie relationship. Seventeen Epimedium spp were classified into two groups of group I and II, since they were divided into two major groups at the similarity coefficients value of 0.65. In addition, one of the two group, group I was divided into three sub-groups including Epimedium koreanum, Chul-won collections 1, 2, and 3, Yanggu, Hongchon, Hwachun, Chunchon, China, Maehwa (Japanese), E. diphyllum and E. violaceum (Japanese), while group II included Chulwon collection 4 and 5 and Japanese collection. The samples collected at Chulwon district showed close similarity with Japanese collection. Similarity indexes between collection and genetic relationship were related at the levels ranging from 0.6 to 0.9.

• Transactions of the Korean hydrogen and new energy society
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• v.17 no.4
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• pp.379-388
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• 2006

Two mesophilic trickling bed bioreactors filled with two different types of media, hydrophilic- and hydrophobic-cubes, were designed and conducted for hydrogen production under the anaerobic fermentation of sucrose. Each bioreactor consisted of the column packed with polymeric cubes and inoculated with heat-treated sludge obtained from anaerobic digestion tank. A defined medium containing sucrose was fed by the different hydraulic retention time(HRT), and recycle rate. Hydrogen concentrations in gas-phase were constant, averaging 40% of biogas throughout the operation. Hydrogen production rate was increased till $10.5\;L{\cdot}h^{-1}{\cdot}L^{-1}$ of bioreactor when influent sucrose concentrations and recycle rates were varied. At the same time, the hydrogen production rate with hydrophobic media application was higher than its hydrophilic media application. No methane was detected when the reactor was under a normal operation. The major fermentation by-products in the liquid effluent of the both trickling biofilters were acetate, butyrate and lactate. In order to run in the long term operation of both reactor filled with hydrophilic and hydrophobic media, biofilm accumulation on hydrophilic media and biogas produced should be controlled through some process such as periodical backwashing or gas-purging. Four sample were collected from each reactor on the opposite hydrogen production rate, and their bacterial communities were compared by terminal restriction fragment length polymorphism (T-RFLP) analysis of PCR products generated using bacterial 16s rRNA gene primers (8f and 926r). It was expressed a marked difference in bacterial communities of both reactors. The trickling bed bioreactor with hydrophobic media demonstrates the feasibility of the process to produce hydrogen gas. A likely application of this reactor technology can be hydrogen gas recovery from pre-treatment of high carbohydrate-containing wastewaters.

• Korean Journal of Clinical Laboratory Science
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• v.48 no.1
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• pp.8-14
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• 2016

Granulosa cells surround the oocyte within the ovarian follicle and play an essential role in creating conditions required for oocyte as well as follicular development. The current study was conducted to examine the gene expression profile of mouse ovaries during the primordial to primary follicle transition process. Total RNAs from mouse ovaries on day 5 and day 12 were synthesized cDNA using annealing control primers. The DEGs were cloned and their identities were analyzed by BLAST search. The Plekha5 and Anxa11 were highly expressed in primary follicle stage. By contrast, their expression was increased in granulosa cells at the primary follicle stage. We have successfully discovered a list of genes expressed in day 5 and day 12 ovaries and confirmed that some of them are differentially expressed in PMF and/or PRI. This is a spatial-temporal regulatory mechanism on the ovarian folliculogenesis through membrane fusion. The gene expression profile from the current study would provide insight for future study on the mechanism(s) involved in primordial-primary follicular transition. This will provide information for identification of the mechanism of ovarian dysfunction.

• The Korean Journal of Mycology
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• v.27 no.2 s.89
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• pp.132-137
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• 1999

This study was carried out to develop the molecular marker for the detection of Pseudomonas tolaasii, a causative agent of bacterial brown blotch disease of oyster mushroom (Pleurotus ostreatus). When several primers designed from repetitive sequences and pectin lyase genes of bacteria were used to produce DNA polymorphism from different Pseudomonas spp. isolated from edible mushrooms, PEU1 primer derived from pectin lyase gene produced polymorphic bands differentiating P. tolaasii strains from other Pseudomonas species. Two bands, 1.0kb and 0.4kb, found commonly in 6 isolates of P. tolaasii were cloned into pGEM-T vector which were designated as pPTOP1 and pPTOP2, respectively, to use as probe. The 0.4 kb insert of pPTOP2 hybridized to only 6 isolates of P. tolaasii, but did not to the other Pseudomonas species. As few as $1.5{\times}10^3$ colony forming unit (cfu) of P. tolaasii could be detected by dot blot hybridization with the cloned 0.4kb DNA in pPTOP2.

• Korean Journal of Microbiology
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• v.40 no.2
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• pp.121-126
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• 2004

Comamonas sp. strain DJ-12 is a 4-chlobiphenyl(4CB)-degrading bacterium that was reidentified from Pseudomonas sp. DJ-12. The genomic DNA was isolated from the strain DJ-12 and amplified by PCR with primers for cloning pcbABCD genes responsible for degradation of 4CB. The amino acid sequences deduced from the nucleotide sequences of pcbA1, pcbA2, pcbA3, pcbA4, pcbB, pcbC2, and pcbD2 genes showed 91, 87, 99, 87, 97, 90 and 87％ homologies with those of Pseudomonas sp. KKS102, respectively. The pcbC1D1 genes that are involved in the degradation of (4-chloro)1,2-dihydroxybiphenyl produced from 4CB by pcbAB gene products were previously reported in the recombinant plasmid pCU1 from Pseudomonas sp. DJ-12. However, the pcbC2D2 genes in the plasmid pCT4 and pCT5 cloned from Comamonas sp. DJ-12 in this study showed 51 and 62％ homologies with those of pcbC1D1 in their nucleotide sequences. The pcbC1D1 and pcbC2D2 genes were found by Southern hybridization to be located at different loci on the chromosome of DJ-12 strain. These results indicate that Comamonas sp. strain DJ-12 has two different sets of pcbCD genes responsible for deg-radation of (4-chloro)1,2-dihydroxybiphenyl.

• Microbiology and Biotechnology Letters
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• v.44 no.3
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• pp.317-323
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• 2016

An isoflavone glucosidase that catalyzes the hydrolysis of isoflavone glucosides into glucose and corresponding aglycones was purified from Candida fermentati SI. The N-terminal sequence was determined to be GLNCDYCN. We designed degenerate primers on the basis of these amino acid sequences and successfully cloned the full structural gene sequence of the isoflavone glucosidase using inverse PCR. The exo-β-(1,3)-glucanase gene consists of 1227 base-pair nucleotides, encoding a 408-amino-acid sequence that shares 41–96% amino acid homology with other yeast exo-β-(1,3)-glucanases belonging to glycoside hydrolase family 5. The recombinant exo-β-(1,3)-glucanase was expressed in Pichia pastoris X-33, using a pPICZA vector system, and further characterized. The molecular mass of the purified exo-β-(1,3)-glucanase was estimated by SDS-PAGE to be 47 kDa. The optimal pH and temperature were pH 4.5 and 40℃, respectively. The K_m values of the purified exo-β-(1,3)-glucanase for daidzin and genistin were 0.12 mM and 0.14 mM, respectively. The V_max values of the purified isoflavone glucosidase were 945.03 U/mg for daidzin and 835.92 U/mg and for genistin.