• Journal of Embryo Transfer
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• v.23 no.3
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• pp.183-187
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• 2008

This study was conducted to analyze the expression pattern of inhibitor of DNA binding proteins (Id)1 and Id2 mRNA on folliculogenesis in rat ovary. The ovaries were obtained from 27 days old Sprague-Dawley rat, fixed, dehydrated, and paraffin embedded. For in situ hybridization, anti-sense and sense Idl and Id2 cRNA probes were prepared and applied to the ovarian section. The ovarian sections were coated with NTB-2 emulsion. After that, the slides were developed and counterstained with hematoxylin and eosin staining. In oocytes, the hybridizational signals of Id1 mRNA were strong in primordial and primary follicles, however, there were no signals in that of atretic or preovulatory follicles. The Id2 mRNA signals were also strong in the oocytes of primordial, primary and secondary follicles. Interestingly, the Id2 mRNA was expressed specifically granulosa cells, but nor in oocyte or theca cells in dominant and preovulatory follicles. Based on these results, Id1 and Id2 mRNA was expressed specifically at follicle stages and follicular tissue and might be closely related with follicle development.

• Journal of Periodontal and Implant Science
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• v.30 no.2
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• pp.389-405
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• 2000

The purpose of this study is to evaluate the effect of IGF-I for DNA synthetic activity and the mRNA expression of bone matrix protein, type I collagen and osteopontin in prolifetation and differentiation of MC3T3-E1 cells. To evaluate DNA synthetic activity, cells were seeded at $2{\times}10^4cells/ml$ in 24 well plates and to evaluate mRNA of type I collagen and osteopontin cells were seeded at $5{\times}10^5cells/ml$ in 100mm culture dishes. These cells were cultured in alpha-minimum essential medium(${\alpha}-MEM$) containing 10% fetal bovine serum at $37^{\circ}C$, 5% $CO_2$ incubator. For DNA synthetic activity test 1, 10, 100ng/ml IGF-I were added to the cells which had been cultured for 3 days before 24 hours. For type I collagen mRNA expression 1, 10ng/ml IGF-I were added to the cells which had been cultured for 5, 10 days and for osteopontin mRNA expression 0.1, 1, 10ng/ml IGF-I were added to the cells which had been cultured for 5, 15, 20 days. Cell proliferaton was measured by the incorporation of [$^3H$]-thymidine into DNA and expression for type I collagen and osteopontin were measured by northern blot analysis. The results were as follows : DNA synthetic activity were generally higher in experimental group than control group. Expressions of type I collagen mRNA were higher at 5 day group and much lower at 10 day group in the control groups. In the experimental groups, mRNA expressions were slightly increased when 1 ng/ml IGF-I were added to 5 day group and decreased in all experimental 10 day groups. Expressions of osteopontin mRNA were higher at 20 day groups and lower at 15 day groups than the control groups. In the experimental groups, mRNA expressions were incereased when 0.1, 1 ng/ml IGF-I were added to 5 day group and in all the 15 day groups, but decreased when 0.1, 1, 10 ng/ml IGF-I were added to 20 day groups. IGF-I stimulated DNA synthetic activity of MC3T3-E1 cells during proliferation stage significantly, did not greatly changed effects on type I collagen mRNA expression and stimulated osteopontin mRNA expression at 15 day especially. In conclusion, we suggests that IGF-I have a tendency of stimulation effect of DNA synthetic activity but do not stimulate type I collagen mRNA in proliferation stage of MC3T3-E1 cell cultures, and stimulate osteopontin mRNA in differentiation stage of MC3T3-E1 cell cultures.

• International Journal of Industrial Entomology and Biomaterials
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• v.6 no.2
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• pp.157-162
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• 2003

The glutathione S-transferase (GSTs) are enzymes responsible for the protection of cells from chemical toxicants and oxidative stress. We describe here the cDNA sequence and mRNA expression of a putative GST from the mole cricket, Gryllotalpa orientalis. The G. orientalis GST cDNA sequences comprised of 621 bp encoding 207 amino acid residues. The multiple sequence alignment of G. orientalis GST gene with other known insect GSTs showed several conserved residues that may be essential for the enzymatic activity of the protein. Phylogenetic analysis of the deduced amino acid sequences of G. orientalis GST gene with other insect GST sequences revealed that the G. orientalis GST gene belongs to class I GST, forming a strong monophyletic group (100% bootstrap value) exclusively for class I GSTs from a diverse insect species. Northern blot analysis confirmed midgut-specific expression at transcriptional level, evidencing the midgut as a site for GST synthesis.

• Journal of Microbiology and Biotechnology
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• v.9 no.4
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• pp.475-481
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• 1999

The study of lignin, a major component of secondary cell wall, has been partly focused on its removal from the woody part in the kraft pulping industry. Cinnamyl alcohol dehydrogenase (CAD; EC 1.1.l95) catalyzes the synthesis of cinnamyl alcohols from corresponding cinnamaldehydes. A cDNA clone, MdCADl, encoding putative CAD from apples (Malus domestica Borkh. cv Fuji) was characterized in this study. The clone contains an open reading frame of 325 amino acid residues, which shows a greater than 80％ identity with Eucalyptus CADl. MdCADl mRNA was detectable in vegetative tissues and was strongly expressed in the fruit. The expression pattern of MdCADl mRNA in the fruit peel after light exposure was also examined. The mRNA was rapidly increased until 1 day after light exposure and remained stable thereafter, suggesting that MdCADl is light inducible. The inducibility of the MdCADl gene was examined using several environmental stresses. Mechanical wounding of leaves increased the MdCADl mRNA level and the induction was further increased by salicylic acid. Southern blot hybridization showed that there is either one or a few copies of CAD genes in apples. To our knowledge, it is believed that MdCADl is the first CAD clone expressed predominantly in fruit.

• KOREAN JOURNAL OF CROP SCIENCE
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• v.38 no.3
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• pp.275-282
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• 1993

The objective of this study was to clone a partial fragment of $\alpha$-amylase from Korean maize. We designed and synthesized an oligonucleotide probe and two kinds of PCR primers based on cDNA conserved region of $\alpha$-amylase sequences from other plants. Total RNA from 3-day-old maize seedling was used as template for 1st strand cDNA synthesis and RNA-DNA hybrid was used as template for polymerase chain reaction(PCR). The product of PCR was about 0.5 kb long and inserted into pUC19. We named this recombinant plasmid as pZM$\alpha$'. The cloned fragment was certified by Southern blot analysis using labeled synthetic oligonucleotide as probe.

• Journal of Life Science
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• v.17 no.3 s.83
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• pp.420-426
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• 2007

In the present investigation, we studied the modulating effects of (-)-epigallocatechin-3-gallate(EGCG) on the differentiation of mouse C2C12 myoblasts. We found that the strong inhibitory effect of EGCG on DNA methyltransferase-mediated DNA methylation induced transdifferentiation of C2C12 myoblasts into smooth muscle cells demonstrated by both morphological changes and immunofluorescent staining. C2C12 myoblasts treated with EGCG for 4 days expressed smooth muscle ${\alpha}-actin$ protein. Real-time PCR data revealed that smooth muscle ${\alpha}-actin$ mRNA was induced by EGCG treated C2C12 myoblasts in a concentration-dependent manner. Smooth muscle ${\alpha}-actin$ mRNA concentration increased 330% and 490% after 2 and 3 days of 50 ${\mu}M$ of EGCG treatment. The expression of another smooth muscle marker, transgelin, mRNA was also increased up to 9-fold by 4 days of EGCG treatment compared with control in a concentration-dependent manner. These results suggested that C2C12 enables to transdifferentiate into smooth muscle when gene expression patterns are changed by the inhibition of DNA methylation induced by EGCG. In conclusion, transdifferentiation of C2C12 myoblasts into smooth muscle is resulted from the modulating effects of EGCG on DNA methylation which subsequently results in changing the expression pattern of several genes playing a critical role in the differentiation of C2C12 myoblasts.

• Plant Resources
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• v.1 no.1
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• pp.13-21
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• 1998

A cDNA Fragment encoding iron storage protrin generated by polymerase chain reaction(PCR) using highly conserved regions of ferritin related genes were used to sereen a red pepper cDNA library. cDNA clone was designated as Fp1. Fp1 clone contatines a 5' nontranslated region of 51dp containing stop conds. Down stream from 5' UTP. an open reading frame of 750bp was observed. followed by a 3' UTR of 272bp. The deduces amino acid sequence of red pepper protein(Fp1) showed 84%, 48% and 36% identity with soybean(SolC). human(HuL H) and horse spleen(HoS-L) ferritin mRNA accumulation in response to iron. Ferritin mRNA accumulation was transient and particularly abundant in leaves. reaching a maxmum at 12h. The level of ferritin mRNA in roots was affected to a lesser extent than in leaves.

• Asian-Australasian Journal of Animal Sciences
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• v.12 no.5
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• pp.695-701
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• 1999

The hypothalamus is the classic site of synthesis of arginine vasotocin as neurohypophyseal hormone in the chicken. However, high concentrations of arginine vasotocin were also measured in ovarian tissues by radioimmunoassay. At first, we observed specific positive signal of mRNA encoding AVT in the hypothalamus by Northern hybridization. However, we could not find any specific bands in ovarian and uterine tissues. For evidence of transcription of the arginine vasotocin gene ingonadal tissues of the chicken, this study has applied the polymerase chain reaction as a highly sensitive assay. The hypothalamus, the four largest preovulatory ovarian follicles and the shell gland (uterus) were collected at 4 h and 20 h before oviposition. The ovarian follicular tissues were separated into granulose theca interns and theca externa layers. The uterine tissues were separated into myometrium and endometrium The extracted mRNA was converted to cDNA by reverse-transcriptase using oligo-$d(T)_{15}$ primer. Then, the cDNA was amplified by Vent polymerase and arginine vasotocin specific primers. The amplification reaction was incubated by 30 cycles successively, $95^{\circ}C$, $55^{\circ}C$ and $72^{\circ}C$ earth for 1 min. Te comparisons of the mRNA levels encoding arginine vasotocin between the tissues were determined by semi-quantification methods. After amplification of the cDNA, the PCR products were detected in hypothalamus, ovarian tissues and uterine tissues. The results of semi-quantification showed that the levels of arginine vasotocin mRNA in ovarian iud uterine tissues were about from 1/50 to 1/1000 when compared to that in the hypothalamus. The very low levels of mRNA encoding arginine vasotocin in ovarian and uterine tissues probably led us to conclude that arginine vasotocin may play a role of local mediate acting autocrine and/or paracrine.

• International Journal of Industrial Entomology and Biomaterials
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• v.7 no.1
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• pp.51-57
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• 2003

A cDNA encoding a putative alcohol dehydrogenase (ADH) class III was cloned from the silkworm, Bombyx mono The full length cDNA is 1,385 nucleotides long and contains an open reading frame of 1,128 bp encoding 376 amino acid residues. The B. mon ADH III protein sequence was aligned with ADH III known from various organisms. Interestingly, the protein sequence of B. mon ADH III showed 87% and 85% identity to ADH III from marine fish Sparus aurata and Branchiostoma floridae, respectively, whereas rather low sequence identity (83%) to Drosophila melanogaster ADH III was observed. Northern blot analysis revealed that B. mon ADH III mRNA is expressed in all tissues from larva examined: fat body, midgut, epidermis, silk gland and ovary, with the highest level found in the fat body.

• International Journal of Industrial Entomology and Biomaterials
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• v.5 no.1
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• pp.103-108
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• 2002

We describe here the cDNA sequence and mRNA expression of a novel serine pretense from the firefly, Pyrocoelia rufa. The 771 bp cDNA encodes for 257 amino acid residues. The deduced protein of P. rufa serine pretense gene contains the catalytic triad and six-conserved cysteine residues. Alignment of the deduced protein of P. rufa serine pretense gene showed 47.4% protein sequence identity to known coleopteran insect Rhyzopertha dominica midgut trpsin-like enzyme. Northern blot analysis revealed that the P. rufa serine pretense is specifically expressed in the midgut of P. rufa larvae.