• Journal of Microbiology and Biotechnology
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• v.21 no.1
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• pp.55-61
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• 2011

Forty yeast strains isolated from soils taken from different locations in Egypt were tested for their P-solubilizing activities on the basis of analyzing the clear zone around colonies growing on a tricalcium phosphate medium after incubation for 5 days at $25^{\circ}C$, denoted as the solubilization index (SI). Nine isolates that exhibited P-solubilization potential with an SI ranging from 1.19 to 2.76 were genetically characterized as five yeasts belonging to the genus Saccharomyces cerevisiae and four non-Saccharomyces, based on a PCR analysis of the ITS1-26S region amplied by SC1/SC2 species-specific primers. The highest P-solubilization efficiency was demonstrated by isolate PSY- 4, which was identified as Saccharomyces cerevisiae by a sequence analysis of the variable D1/D2 domain of the 26S rDNA. The effects of single and mixed inoculations with yeast PSY-4 and Bacillus polymyxa on the P-uptake and growth of corn were tested in a greenhouse experiment using different levels of a phosphorus chemical fertilizer (50, 100, and 200 kg/ha super phosphate 15.5% $P_2O_5$). The results showed that inoculating the corn with yeast PSY-4 or B. polymyxa caused significant increases in the shoot and root dry weights and P-uptake in the shoots and roots. The P-fertilization level also had a significant influence on the shoot and root dry weights and P-uptake in the shoots and roots when increasing the P-level from 50 up to 200 kg/ha. Dual inoculation with yeast strain PSY-4 and B. polymyxa at a P-fertilization level of 200 kg/ha gave higher values for the shoot and root dry weights and P-uptake in the shoots and roots, yet these increases were nonsignificant when compared with dual inoculation with yeast strain PSY-4 and B. polymyxa at a P-fertilization level of 100 kg/ha. The best increases were obtained from dual inoculation with yeast strain PSY-4 and B. polymyxa at a P-fertilization level of 100 kg/ha, which induced the following percentage increases in the shoot and root dry weights, and P-uptake in the shoots and roots; 16.22%, 46.92%, 10.09%, and 31.07%, respectively, when compared with the uninoculated control (fertilized with 100 kg/ha).

• Journal of Plant Biotechnology
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• v.42 no.4
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• pp.356-363
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• 2015

In order to study genetic engineering in trees, the characterization of genes and promoters from trees is necessary. We isolated the promoter region (867 bp) of Pagns-LTP from poplar (P. alba ${\times}$ P. glandulosa) and characterized its activity in transgenic poplar plants using a ${\beta}$-glucuronidase (GUS) reporter gene. High-level expression of the Pagns-LTP transcript was found in poplar roots, while comparatively low-level expression was found in the young leaves. Pagns-LTP mRNA was not detected in other poplar tissues. Additionally, transgenic poplar plants that contained a Pagns-LTP promoter fused to a GUS reporter gene, displayed tissue-specific GUS enzyme activity localized in root tissue. In silico analysis of the Pagns-LTP promoter sequence reveals the presence of several cis-regulatory elements responsive to phytohormones, biotic and abiotic stresses, as well as those regulating tissue-specific expression. These results demonstrate that the Pagns-LTP promoter has tissue-specific expression activity in poplar roots and leaves that may be involved in organ development and plant resistance to various stresses. Therefore, we anticipate that the Pagns-LTP promoter would be a useful tool to genetically optimize woody plants for functional genomics.

• Korean Journal of Microbiology
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• v.28 no.2
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• pp.91-97
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• 1990

Complementary DNA (cDNA) coding for human cytoplasmic superoxide dismutase (SOD1) (superoxide: superoxide oxidoreductase E.C.1.15.1.1) was isolated from human liver cDNA library of $\lambda$gt11 by in situ plaque hybridization. The insery cDNA gas the 5' untranslational region (UTR) and 3'UTR of SOD1 gene. Polymerase Chain Reaction (PCR) method was used fro subcloning of SOD1 structural gene. Using synthetic sense strand primer (24mer) containing a start codon and antisense strand primer (24mer), SOD1 structural gene was selectively amplified. Amplified DNA was directly cloned into the HincII site of pUC19 plasmid. Insery cDNA was subcloned into M13 mp19 and sequenced by dideowy chain termination method with Sequenase. The nucleotide sequence of insert cDNA had an open reading frame (ORF) coding for 153 amino acid residues. The structural gene of cytoplasmic SOD was placed under the control of bacteriophage $\lambda P_{L}$ regulatory sequences, generating a highly efficient expression plasmid. The production of human SOD1 in E. coli cells was about 7% of total cellular proteins and recombinant human SOD1 possessed its own enzymatic acitivity.

• BMB Reports
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• v.38 no.1
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• pp.58-64
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• 2005

Myo-inositol monophosphate phosphatase (IMPP) is a key enzyme in the phosphoinositide cell-signaling system. This study found that incubating the IMPP from a porcine brain with pyridoxal-5'-phosphate (PLP) resulted in a time-dependent enzymatic inactivation. Spectral evidence showed that the inactivation proceeds via the formation of a Schiff's base with the amino groups of the enzyme. After the sodium borohydride reduction of the inactivated enzyme, it was observed that 1.8 mol phosphopyridoxyl residues per mole of the enzyme dimer were incorporated. The substrate, myo-inositol-1-phosphate, protected the enzyme against inactivation by PLP. After tryptic digestion of the enzyme modified with PLP, a radioactive peptide absorbing at 210 nm was isolated by reverse-phase HPLC. Amino acid sequencing of the peptide identified a portion of the PLP-binding site as being the region containing the sequence L-Q-V-S-Q-Q-E-D-I-T-X, where X indicates that phenylthiohydantoin amino acid could not be assigned. However, the result of amino acid composition of the peptide indicated that the missing residue could be designated as a phosphopyridoxyl lysine. This suggests that the catalytic function of IMPP is modulated by the binding of PLP to a specific lysyl residue at or near its substrate-binding site of the protein.

• Microbiology and Biotechnology Letters
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• v.31 no.4
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• pp.322-328
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• 2003

The Transformation-Associated Recombination (TAR) cloning technique allows selective isolation of chromosomal regions and genes from complex genomes. The procedure requires knowledge of relatively small genomic sequences that reside adjacent to the chromosomal region of interest. This technique involves homologous recombination during yeast spheroplast transformation between genomic DNA and a TAR vector that has 5'and 3' gene targeting sequences. In this study, we examined the minimum size of specific hooks required for a single-copy gene isolation and compared the utility of different TAR vectors, radial and unique vectors, by cloning the same single-copy gene. The efficiency of TAR cloning of the hHPRT gene was same using hooks varying from 750 to 63 bp. The number of transformants decreased approximately 20-fold when the TAR vector contained two unique hooks versus using a radial vector, but the percentage of positive recombinants increased over 2-fold when a unique TAR vector was used. Therefore, we suggest that the two-unique TAR vector is suitable for general TAR cloning given its high selectivity, and the radial TAR vector is more suitable when genomic DNA is in limited quantity, for example, DNA isolated from pathological specimens. Moreover, we confirm the minimal length of a unique sequence in a TAR vector is approximately 60 bp for a single-copy gene isolation.

• Asian-Australasian Journal of Animal Sciences
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• v.22 no.8
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• pp.1078-1084
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• 2009

The Jindo dog is a Korean natural monument and is recognized by the Fédération Cynologique Internationale. A prominent feature is the diverse coat color within the breed. To analyze the genetic basis of variation in the Jindo coat color, we sequenced the protein-coding regions of the melanocortin 1 receptor gene (MC1R). The MC1R coding sequence was determined from 154 dogs in five breeds (Jindo, Labrador Retriever, English Springer Spaniel, Belgian Malinois, and German Shepherd). To confirm the genetic structure of sampled populations, we tested for Hardy-Weinberg equilibrium (HWE) and computed $F_{st}$ The sample populations did not significantly deviate from HWE. $F_{st}$ was 0.02 between white and fawn Jindo dogs; this was lower than $F_{st}$ between breeds. Six single nucleotide polymorphisms (SNPs) were detected in the MC1R coding region. Among the six SNPs, five were non-synonymous (S90G, T105A, Q159P, M264V, and R306ter) and one was synonymous SNP (Y298Y). From the SNPs, we predicted four haplotypes (H1, H2, H3, and H4) for Jindo MC1R. Jindo dogs had different haplotypes corresponding to different coat colors. H1 was frequently observed in white Jindo dogs with an odds ratio of 5.03 (95% CI: 2.27-11.18, p<0.0001), whereas H2 and H4 were observed only in fawn Jindo dogs. Our findings indicate that SNP haplotype can influence coat color. Knowledge of MC1R haplotypes can help discriminate white and fawn coats in Jindo dogs. We hope this report will trigger more research into the genetics of this traditional Korean dog and will be a reference for dogs of Asian origin. Also, our results will provide a useful genetic marker for Jindo dog breeders who have selected for specific colors.

• The Korean Journal of Physiology and Pharmacology
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• v.9 no.6
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• pp.333-339
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• 2005

B/K protein is a novel protein containing double C2-like domains. We examined the specific signaling pathway that regulates the transcription of B/K in PC12 cells. When the cells were treated with forskolin ($50{\mu}M$), B/K mRNA and protein levels were time-dependently decreased, reaching the lowest level at 3 or 4 hr, and thereafter returning to the control level. Chemicals such as dibutyryl-cAMP, cellpermeable cyclic AMP (cAMP) analogue and CGS21680, adenosine receptor $A_{2A}$ agonist, also repressed the B/K transcription. However, 1,9-dideoxyforskolin did not show inhibitory effect on B/K transcription, suggesting direct involvement of cAMP in the forskolin-induced inhibition of B/K transcription. Effect of forskolin, dibutyryl cAMP and CGS21680 was significantly reduced in PKA-deficient PC12 cell line (PC12-123.7). One cAMP-response element (CRE)-like sequence (B/K CLS) was found in the promoter region of B/K DNA, and electrophoretic mobility shift assay indicated its binding to CREM and CREB. Forskolin significantly suppressed the promoter activity in CHO-K1 cells transfected with the constructs containing B/K CLS, but not with the construct in which B/K CLS was mutated (AC：TG). Taken together, we suggest that the transcription of B/K gene in PC12 cells may be regulated by PKA-dependent mechanism.

• Journal of the Korean Institute of Intelligent Systems
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• v.15 no.2
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• pp.163-172
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• 2005

When tracking a human face in the moving pictures with complex background under irregular lighting conditions, the detected face can be larger including background or smaller including only a part of the face. Even background can be detected as a face area. To solve these problems, this paper proposes a new face tracking method using a block difference image and a Kalman estimator. The block difference image allows us to detect even a small motion of a human and the face area is selected using the skin color inside the detected motion area. If the pixels with skin color inside the detected motion area, the boundary of the area is represented by a code sequence using the 8-neighbor window and the head area is detected analysing this code. The pixels in the head area is segmented by colors and the region most similar with the skin color is considered as a face area. The detected face area is represented by a rectangle including the area and its four vertices are used as the states of the Kalman estimator to trace the motion of the face area. It is proved by the experiments that the proposed method increases the accuracy of face detection and reduces the fare detection time significantly.

• Asian Pacific Journal of Cancer Prevention
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• v.15 no.18
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• pp.7589-7595
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• 2014

Apurinic/apyrimidinic endonuclease 1 (APEX1) is a multifunctional protein which plays a central role in the BER pathway. APEX1 gene being highly polymorphic in cancer patients and has been indicated to have a contributive role in Apurinic/apyrimidinic (AP) site accumulation in DNA and consequently an increased risk of cancer development. In this case-control study, all exons of the APEX1 gene and its exon/intron boundaries were amplified in 530 breast cancer patients and 395 matched healthy controls and then analyzed by single-stranded conformational polymorphism followed by sequencing. Sequence analysis revealed fourteen heterozygous mutations, seven 5'UTR, one 3'UTR, two intronic and four missense. Among identified mutations one 5'UTR (rs41561214), one 3'UTR (rs17112002) and one missense mutation (Ser129Arg, Mahjabeen et al., 2013) had already been reported while the remaining eleven mutations. Six novel mutations (g.20923366T>G, g.20923435G>A, g.20923462G>A, g.20923516G>A, 20923539G>A, g.20923529C>T) were observed in 5'UTR region, two (g.20923585T>G, g.20923589T>G) in intron1 and three missense (Glu101Lys, Ala121Pro, Ser123Trp) in exon 4. Frequencues of 5'UTR mutations; g.20923366T>G, g.20923435G>A and 3'UTR (rs17112002) were calculated as 0.13, 0.1 and 0.1 respectively. Whereas, the frequency of missense mutations Glu101Lys, Ser123Trp and Ser129Arg was calculated as 0.05. A significant association was observed between APEX1 mutations and increased breast cancer by ~9 fold (OR=8.68, 95%CI=2.64 to 28.5) with g.20923435G>A (5'UTR), ~13 fold (OR= 12.6, 95%CI=3.01 to 53.0) with g.20923539G>A (5'UTR) and~5 fold increase with three missense mutations [Glu101Lys (OR=4.82, 95%CI=1.97 to 11.80), Ser123Trp (OR=4.62, 95%CI=1.7 to 12.19), Ser129Arg (OR=4.86, 95%CI=1.43 to 16.53)]. The incidence of observed mutations was found higher in patients with family history and with early menopause. In conclusion, our study demonstrates a significant association between germ line APEX1 mutations and breast cancer patients in the Pakistani population.

• Asian-Australasian Journal of Animal Sciences
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• v.30 no.10
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• pp.1471-1477
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• 2017

Objective: Since athletic performance is a most importance trait in horses, most research focused on physiological and physical studies of horse athletic abilities. In contrast, the molecular analysis as well as the regulatory pathway studies remain insufficient for evaluation and prediction of horse athletic abilities. In our previous study, we identified AXL receptor tyrosine kinase (AXL) gene which was expressed as alternative spliced isoforms in skeletal muscle during exercise. In the present study, we validated two AXL alternative splicing transcripts (named as AXLa for long form and AXLb for short form) in equine skeletal muscle to gain insight(s) into the role of each alternative transcript during exercise. Methods: We validated two isoforms of AXL transcripts in horse tissues by reverse transcriptase polymerase chain reaction (RT-PCR), and then cloned the transcripts to confirm the alternative locus and its sequences. Additionally, we examined the expression patterns of AXLa and AXLb transcripts in horse tissues by quantitative RT-PCR (qRT-PCR). Results: Both of AXLa and AXLb transcripts were expressed in horse skeletal muscle and the expression levels were significantly increased after exercise. The sequencing analysis showed that there was an alternative splicing event at exon 11 between AXLa and AXLb transcripts. 3-dimentional (3D) prediction of the alternative protein structures revealed that the structural distance of the connective region between fibronectin type 3 (FN3) and immunoglobin (Ig) domain was different between two alternative isoforms. Conclusion: It is assumed that the expression patterns of AXLa and AXLb transcripts would be involved in regulation of exercise-induced stress in horse muscle possibly through an $NF-{\kappa}B$ signaling pathway. Further study is necessary to uncover biological function(s) and significance of the alternative splicing isoforms in race horse skeletal muscle.