• BMB Reports
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• v.42 no.8
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• pp.493-499
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• 2009

MicroRNAs (miRs) are a family of small noncoding RNAs that regulate gene expression by targeting mRNAs in a sequence specific manner, inducing translational repression or mRNA degradation. In this paper we have first analyzed by microarray the miR-profile in erythroid precursor cells from one normal and two thalassemic patients expressing different levels of fetal hemoglobin (one of them displaying HPFH phenotype). The microarray data were confirmed by RT-PCR analysis, and allowed us to identify miR-210 as an highly expressed miR in the erythroid precursor cells from the HPFH patient. When RT-PCR was performed on mithramycin-induced K562 cells and erythroid precursor cells, miR-210 was found to be induced in time-dependent and dose-dependent fashion, together with increased expression of the fetal $\gamma$-globin genes. Altogether, the data suggest that miR-210 might be involved in increased expression of $\gamma$-globin genes in differentiating erythroid cells.

• Journal of Life Science
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• v.8 no.3
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• pp.249-256
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• 1998

The purpose of this study was to understand the evolutionary relationships between fish and other vertebrates which had DNA with the genetic defects in homoglobin expression, with comparison to the nucleotide homologies of the ${\beta}$-globin genes. The predicted amino acid sequence from carp ${\beta}$-globin gene was compared with those of other vertebrates from the published data. The nucleotide homologies of the predicted amino acid sequence from the carp ${\beta}$-globin gene with those of goldfish and mirror carp were high, and the rates were 97.3% and 93.9%, respectively. On the other hand, with the previously reported ${\beta}$-globins of goat, frog, human, rat, goose, chicken, and duck, it showed low homology ranging from 45.9 to 58.1%. The carp ${\beta}$-globin has one inserted amino acid residue, which was also found in other fish ${\beta}$globin, but not in other vertebrate ${\beta}$-globins.

• Journal of Life Science
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• v.22 no.11
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• pp.1587-1594
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• 2012

3C (chromatin conformation capture) is a technique to analyze chromatin organization in nuclei of eukaryotic cells. The procedure of 3C includes the formaldehyde treatment of cells to fix interactions between proteins and between proteins and DNA in chromatin, the digestion of fixed chromatin with restriction enzyme, and the ligation of fragmented DNA. The efficiency of DNA ligation represents proximity between DNA fragments in chromatin organization. Studies in the ${\beta}$-globin locus using 3C showed that the locus control region is in close proximity to the transcriptionally-active globin genes, indicating that chromatin organization has a role in transcriptional regulation of the genes. 3C has been advanced by combining with ChIP and genome-wide sequencing. This review presents the principle and procedure of the 3C technique, the chromatin organization of the ${\beta}$-globin locus explained by 3C, and advanced techniques based on 3C.

• Journal of Microbiology and Biotechnology
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• v.15 no.3
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• pp.544-550
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• 2005

To develop a highly efficient mammalian expression vector, a series of vectors were constructed based on the murine cytomegalovirus (MCMV) immediate-early (IE) promoter and human ${\beta}$-globin second intron. The resulting MCMV promoter was several-fold stronger than the HCMV promoter in various mammalian cell lines, such as the NIH3T3, Neuro-2a, 293T, and HT1080 cell lines, and was only slightly weaker than the HCMV promoter in HeLa and CHO cells. The inclusion of the human ${\beta}$-globin second intron behind the MCMV promoter or HCMV promoter markedly enhanced the promoter activity in various mammalian cell lines, and the resultant MCMV/Glo-I expression system was stronger than the HCMV promoter from 4.7- to 11.2-fold in every cell line tested. Also, the MCMV/Glo-I promoter induced a higher level of the VSV-G protein in a transiently transfected 293T cell line, which is useful for the production of recombinant retrovirus and lentivirus vectors.

• Reproductive and Developmental Biology
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• v.31 no.4
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• pp.241-248
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• 2007

Epigenetic modification dependent DNA methyltransferases (DNMTs) play an important role in tissue- and stage-specific gene regulation and normal mammalian development. In this study, we show that DNMTs are expressed at different levels during hematopoietic stem cell (HSC) differentiation to proerythrocytes. DNMT1, DNMT3A, and DNMT3B were highly expressed at day 7 after differentiation. We used specific siRNA as a tool to probe the relationship between the expression of DNMTs and erythropoietic differentiation. When introduced siRNA of DMNT1 and DMNT3b in human $CD34^+$ cells, these more differentiated into erythrocytes. This was confirmed by glycophorin A (GPA) positive cell analysis and globin gene expression. $GPA^+$ cells increased up to $20{\sim}30%$, and ${\gamma}$- and ${\epsilon}$-globin genes increased in siRNA transfected cells. Therefore, our data suggest that suppression of DNA methylation can affect positively differentiation of HSC and may contribute to expression of erythrocyte lineage genes including GPA and globins.

• BMB Reports
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• v.37 no.2
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• pp.139-143
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• 2004

The antioxidant responsive element (ARE) is a cis-acting regulatory element of genes encoding phase II detoxification enzymes and antioxidant proteins, such as NAD(P)H: quinone oxidoreductase 1, glutathione S-transferases, and glutamate-cysteine ligase. Interestingly, it has been reported that Nrf2 (NF-E2-related factor 2) regulates a wide array of ARE-driven genes in various cell types. Nrf2 is a basic leucine zipper transcription factor, which was originally identified as a binding protein of locus control region of ss-globin gene. The DNA binding sequence of Nrf2 and ARE sequence are very similar, and many studies demonstrated that Nrf2 binds to the ARE sites leading to up-regulation of downstream genes. The function of Nrf2 and its downstream target genes suggests that the Nrf2-ARE pathway is important in the cellular antioxidant defense system. In support of this, many studies showed a critical role of Nrf2 in cellular protection and anti-carcinogenicity, implying that the Nrf2-ARE pathway may serve as a therapeutic target for neurodegenerative diseases and cancers, in which oxidative stress is closely implicated.

• Molecules and Cells
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• v.20 no.1
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• pp.57-68
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• 2005

Murine erythroleukemia (MEL) cells are widely used to study erythroid differentiation thanks to their ability to terminally differentiate in vitro in response to chemical induction. At the molecular level, not much is known of their terminal differentiation apart from activation of adult-type globin gene expression. We examined changes in gene expression during the terminal differentiation of these cells using microarray-based technology. We identified 180 genes whose expression changed significantly during differentiation. The microarray data were analyzed by hierarchical and k-means clustering and confirmed by semi-quantitative RT-PCR. We identified several genes including H1f0, Bnip3, Mgl2, ST7L, and Cbll1 that could be useful markers for erythropoiesis. These genetic markers should be a valuable resource both as potential regulators in functional studies of erythroid differentiation, and as straightforward cell type markers.

• The Korean Journal of Zoology
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• v.39 no.3
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• pp.239-247
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• 1996

An embryonic stem (ES) cell-based in vitro model system was examined to determine whether a simple differentiation of embryoid bodies (EB) in the suspension medium is useful to dissect early erythropoiesis. Characteristics of the differentiating EBs were monitored for their differentiation potential to generate hematopoietic cell types by general morphology, benzidine staining and two-step colony assays, and expressivity of several erythroid marker genes by the RT-PCR analysis for total cellular RNA prepared from the differentiating EBs. Every ematopoietic lineage cells were generated from the differentiating EBs with reproducible frequencies, similar to the other sophisticated differentiation protocols. Furthermore, the globin gene switching in differentiating ES cells paralleled the sequence of events found in the mouse embryo, and such that their expression was activated by at least 12 hrs later than those of erythroid-specific transcription factors, GATA-1 and Tal-1 The erythropoietic differentiation program initiated reproducibly and efficiently in this simple differentiation system in a suspension culture, such that this system may be useful for dissection of the molecular events of early erythropoiesis.

• BMB Reports
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• v.39 no.3
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• pp.292-296
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• 2006

DNA sequencing has resulted in an abundance of data on DNA sequences for various species. Hence, the characterization and comparison of sequences become more important but still difficult tasks. In this paper, we first give a 2-D ladderlike graphical representation for the characteristic sequences of a DNA sequence, and then construct a 3-component vector, in which the normalized ALE-indices extracted from such three 2-D graphs via D/D matrices are individual components, to characterize the DNA sequence. The examination of similarities/dissimilarities among sequences of the $\beta$-globin genes of different species illustrates the utility of the approach.

• Asian Pacific Journal of Cancer Prevention
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• v.14 no.4
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• pp.2355-2359
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• 2013

Vulva cancer is rare among all gynecological cancer worldwide, including Thailand, and mainly affects older women. Persistent high risk type infection of human papillomavirus (HPV) is the one important factor for developing cancer. In this study, we focused on HPV DNA investigation and type-specific distribution of HPV in 25 formalin-fixed paraffin-embedded (FFPE) samples collected from Thai women with vulva cancer histologically confirmed by the National Cancer Institute, Thailand, during 2003-2011. HPV DNA detection and genotyping were undertaken with polymerase-chain reaction and enzyme-immunoassay using GP5+/bio6+ consensus specific primers and digoxigenin-labeled specific oligoprobes, respectively. Human ${\beta}$-globin genes was used as the internal control. Our results showed that 44% (11/25) of all vulva cancer samples were HPV-positive. All of them are high risk HPV type infection, detected as single (63.64%, 7/11) and/or double infections (4/11, 36.36%). HPV 16 was the most common type identified in vulva cancer, followed by HPV 35, 33, 18 and 58. In conclusion, this study presented that HPV-16 is observed at the highest frequency in this cancer, similar to cervical cancer, with HPV 18 being less frequent. Although the sample size was small and could not represent overall incidence and prevalence in Thai women, these preliminary data for vulva cancer are of interest since they reinforce the necessity for HPV screening or vaccination in Thailand.