Optimized Transformation of Streptomyces sp. ATCC 39366 Producing Leptomycin by Electroporation

Yong-Qiang Fan,Hong-Jian Liu,Li Yan,Yu-Shi Luan,Hai-Meng Zhou,Jun-Mo Yang,Shang-Jun Yin,Yu-Long Wang 한국미생물학회 2013 The journal of microbiology Vol.51 No.3

Streptomyces sp. ATCC 39366 produces leptomycin derivatives. Leptomycin B, a potent and specific inhibitor against the export of nuclear proteins, is the main product; however, the introduction of DNA into this strain is almost impossible, which has impeded its further use. We developed a Streptomyces sp. ATCC 39366 transformation protocol to introduce foreign DNA via electroporation. Various conditions were examined, including treatments of the cell wall with weakening agents, electroporation parameters, and DNA content. We found that only plasmid DNA isolated from a dam- ET12567 strain resulted in successful transformation. The mycelium growing in a yeast-peptone-dextrose medium supplemented with 1% glycine at 28°C on a rotary shaker (220 rpm) was more dispersed than those without supplementation and prone to electroporation. The maximum transformation efficiency of 8×102 CFU/μg plasmid DNA was obtained at a field strength of 13 kV/cm with a time constant of 13 ms (25-μF capacitor; parallel resistance, 600 Ω) using 1-mm electrocuvettes. The results of the transformations of two other Streptomyces species indicated that the optimized conditions established in this study might only be applicable to Streptomyces sp. ATCC 39366. However, this is the first report of successful transformation of Streptomyces sp. ATCC 39366, and will facilitate the construction of a gene knockout mutant in Streptomyces sp. ATCC 39366 to produce series of new leptomycin derivatives.

Inactivation of recombinant human brain-type creatine kinase during denaturation by guanidine hydrochloride in a macromolecular crowding system.

Fan, Yong-Qiang,Liu, Hong-Jian,Li, Chang,Luan, Yu-Shi,Yang, Jun-Mo,Wang, Yu-Long Humana Press 2013 Applied biochemistry and biotechnology Vol.169 No.1

<P>In this study, we quantitatively examined the effects of the macromolecular crowding agents, polyethylene glycol 2000 (PEG 2000) and dextran 70, on guanidine hydrochloride (GdnHCl)-induced denaturation of recombinant human brain-type creatine kinase (rHBCK). Our results showed that both PEG 2000 and dextran 70 had a protective effect on the inactivation of rHBCK induced by 0.5 M GdnHCl at 25 C. The presence of 200 g/L PEG 2000 resulted in the retention of 35.33 % of rHBCK activity after 4 h of inactivation, while no rHBCK activity was observed after denaturation in the absence of macromolecular crowding agents. The presence of PEG 2000 and dextran 70 at a concentration of 100 g/L could decelerate the k (2) value of the slow track to 21 and 33 %, respectively, in comparison to values obtained in the absence of crowding agents. Interestingly, inactivation of rHBCK in the presence of 200 g/L PEG 2000 followed first-order monophasic kinetics, with an apparent rate constant of 8??10(-5)?s(-1). The intrinsic fluorescence results showed that PEG 2000 was better than dextran 70 at stabilizing rHBCK conformation. In addition, the results of the phase diagram indicate that more intermediates may be captured when rHBCK is denatured in a macromolecular crowding system. Mixed crowding agents did not produce better results than single crowding agents, but the protective effects of PEG 2000 on the inactivation and unfolding of rHBCK tended to increase as the ratio of PEG 2000 increased in the mixed crowding agent solution. Though it is not clear which crowding agents more accurately simulated the intracellular environment, this study could lead to a better understanding of protein unfolding in the intracellular environment.</P>

Correlation between Non-Alcoholic Fatty Liver Disease and Visceral Adipose Tissue in Non-Obese Chinese Adults: A CT Evaluation

Ai-Hong Yu,Yang-Yang Duan-Mu,Yong Zhang,Ling Wang,Zhe Guo,Yong-Qiang Yu,Yu-Sheng Wang,Xiao-Guang Cheng 대한영상의학회 2018 Korean Journal of Radiology Vol.19 No.5

Objective: To investigate the correlation between non-alcoholic fatty liver disease and visceral adipose tissue in non-obese Chinese adults using computed tomography (CT). Materials and Methods: The study included 454 subjects undergoing abdominal CT scan. Degree of CT attenuation in liver and spleen, and the degree of fat infiltration in liver were evaluated according to three indices: the attenuation value of liver parenchyma (CTLP), the attenuation ratio of liver and spleen (LSratio) and the attenuation difference between liver and spleen (LSdif). Visceral fat area (VFA) and total fat area (TFA) at L2/3 and L4/5 levels were measured, and the abdominal subcutaneous fat area (SFA) was calculated. Bivariate correlation analysis was carried out to determine the correlation among these factors. Results: In men, VFA, SFA and TFA at L2/3 and L4/5 levels showed significant differences in terms of the three indices to distinguish fatty liver from non-fatty liver (all, p < 0.001). In men, all the three indices showed negative correlation with TFA, SFA and VFA (all, p < 0.001). The negative correlation between the three indices and VFA at the L2/3 level was higher than at L4/5 level (r = -0.476 vs. r = -0.340 for CTLP, r = -0.502 vs. r = -0.413 for LSratio, r = -0.543 vs. r = -0.422 for LSdif, p < 0.001, respectively). The negative correlation between LSratio, LSdif and VFA at L2/3 and L4/5 levels was higher than SFA at the corresponding level. In women, all the three indices showed negative correlation with VFA and TFA at L2/3 and L4/5 levels, and the negative correlation between CTLP and VFA was higher at L2/3 level than at L4/5 level (r = -0.294 vs. r = -0.254, p < 0.001). Conclusion: In non-obese Chinese adults, the degree of hepatic fatty infiltration showed a strong correlation with abdominal fat on CT. VFA at L2/3 level was more closely related to fatty liver compared with VFA at L4/5 level.

A chloroplast variation map generated using whole genome re-sequencing of Korean landrace rice reveals phylogenetic relationships among Oryza sativa subspecies

Wei Tong,Qiang He,Xiao-Qiang Wang,Min-Young Yoon,Won-Hee Ra,Feng Peng Li,Jie Yu,Win Htet Oo,Sun-Kyung Min,Buung Choi,Eun-Beom Heo,Byoung-Kook Yun,Kyu-Won Kim,Tae-Sung Kim,Chang-Yong Lee,Yong-Jin Park 한국육종학회 2015 한국육종학회 심포지엄 Vol.2015 No.07

Although the overall structure of the chloroplast genome is generally conserved, a number of sequence variations have been identified, which are valuable for plant population and evolutionary studies. Here, we constructed a chloroplast variation map of 30 landrace rice strains of Korean origin, using the Oryza rufipogon chloroplast genome (Genbank: NC_017835) as a reference. Differential distribution of single nucleotide polymorphisms (SNPs) and indels across the rice chloroplast genome is suggestive of a region-specific variation. Population structure clustering revealed the existence of two clear subgroups (indica and japonica) and an admixture group (aus). Phylogenetic analysis of the 30 landrace rice strains and six rice chloroplast references suggested and supported independent evolution of O. sativa indica and japonica. Interestingly, two “aus” type accessions, which were thought to be indica type, shared a closer relationship with the japonica type. One hypothesis is that “Korean aus” was intentionally introduced and may have obtained japonica chloroplasts during cultivation. We also calculated the nucleotide diversity of 30 accessions and compared to six rice chloroplast references, which shown a higher diversity in the indica and aus groups than in the japonica group in lower level substitution diversity.

Overexpression of Cyclooxygenase-1 Correlates with Poor Prognosis in Renal Cell Carcinoma

Yu, Zu-Hu,Zhang, Qiang,Wang, Ya-Dong,Chen, Jing,Jiang, Zhi-Mao,Shi, Min,Guo, Xin,Qin, Jie,Cui, Guang-Hui,Cai, Zhi-Ming,Gui, Yao-Ting,Lai, Yong-Qing Asian Pacific Journal of Cancer Prevention 2013 Asian Pacific journal of cancer prevention Vol.14 No.6

The aim of this study was to evaluate expression of COX-1 in renal cell carcinoma (RCC) and its prognostic value. mRNA of COX-1 was detected in 42 paired RCC and adjacent normal tissues with quantitative realtime polymerase chain reaction (qRT-PCR). Expression of COX-1 was also evaluated in 196 RCC sections and 91 adjacent normal tissues with immunohistochemistry. Statistical analysis was performed to assess COX-1 expression in RCC and its prognostic significance. The results of qRT-PCR showed mRNA levels of COX-1 in RCC tissues to be significantly higher than that in adjacent normal tissues (p < 0.001). Immunohistochemical assays also revealed COX-1 to be overexpressed in RCC tissues (p < 0.001). Statistical analysis demonstrated high expression of COX-1 was correlated with tumour size (p = 0.002), pathological stage (p = 0.003), TNM stage (p = 0.003, 0.007, 0.027, respectively), and tumour recurrence (p < 0.001). Survival analysis indicated patients with high expression of COX-1 had shorter survival time (p < 0.001), and COX-1 was an independent predictor. This is the first study to reveal overexpression of COX-1 in RRC and point to use as a prognostic marker in affected patients.

Resequencing reveals different domestication rate for BADH1 and BADH2 in rice (Oryza sativa)

Qiang He,Jie Yu,Tae-Sung Kim,Yoo-Hyun Cho,Young-Sang Lee,Yong-Jin Park 한국육종학회 2015 한국육종학회 심포지엄 Vol.2015 No.07

BADH1 and BADH2 are two homology genes, encoding betaine aldehyde dehydrogenase in rice. In the present study, we scanned BADHs sequences of 295 rice cultivars, and 10 wild rice accessions to determine the polymorphisms, gene functions and domestication of these two genes. A total of 16 alleles for BADH1 and 10 alleles for BADH2 were detected in transcription region of cultivars and wild species. Association study showed that BADH1 has significant correlation with salt tolerance in rice during germination stage, the SNP P11483(T/A)ishighlycorrelatedwithsalttoleranceindex(STI)(P<10-4). While, BADH2 was only responsible for rice fragrance, of which two BADH2 alleles (P23036, P25390) explain 97% of aroma variation in our germplasm. It indicated that there are no overlapping functions between the two homology genes. In addition, a large LD block was detected in BADH2 region, however, no large LD blocks in a 4-Mb region of BADH1. Only BADH2 region shown significant bias Tajima’s D value from the balance. Extended haplotype homozygosity study revealed fragrant accessions had a large LD block that extended around the mutation site (P23036) of BADH2, while both of the BADH1 alleles (SNP P11483(T/A)) did not show large extended LDblock. All these results suggested that BADH2 was identified as a domesticated gene during rice evolution, while BADH1 was not selected by human beings.

Genotypic Variation in Fatty Acids in Whole Grain Sesame

Qiang He,Taerim Lee,Jie Yu,Win Htet Oo,Min-Young Yoon,Myeong-Hyeon Min,Sang-Ho Chu,Kyu-Won Kim,Young-Sang Lee,Yong-Jin Park 한국작물학회 2020 Journal of crop science and biotechnology Vol.23 No.1

Fatty acids (FAs) are important desirable compounds in oil crops. Here, we evaluated the amounts of 14 FAs in sesame grains using gas chromatography mass spectrometry and investigated the effects of genotype and environment on FA composition in 2 years. Unsaturated FA fractions accounted for 80.018% of the total FAs in sesame in 2014 and 78.936% in 2015. C16:1 and C20:1 levels exhibited few effects from the environment, while other FAs were affected significantly by environmental variance. C18:1 and C24:0 were significantly correlated with population structure. We also identified 249 QTLs that were strongly associated with the amounts of 14 FAs in sesame. These findings will be useful for understanding FA metabolism in sesame and for improving the quality of sesame through molecular breeding in the future.

A Genome-Wide Association Study Reveals Candidate Genes Related to Salt Tolerance in Rice ( <i>Oryza sativa</i> ) at the Germination Stage

Yu, Jie,Zhao, Weiguo,Tong, Wei,He, Qiang,Yoon, Min-Young,Li, Feng-Peng,Choi, Buung,Heo, Eun-Beom,Kim, Kyu-Won,Park, Yong-Jin MDPI 2018 INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES Vol.19 No.10

<P>Salt toxicity is the major factor limiting crop productivity in saline soils. In this paper, 295 accessions including a heuristic core set (137 accessions) and 158 bred varieties were re-sequenced and ~1.65 million SNPs/indels were used to perform a genome-wide association study (GWAS) of salt-tolerance-related phenotypes in rice during the germination stage. A total of 12 associated peaks distributed on seven chromosomes using a compressed mixed linear model were detected. Determined by linkage disequilibrium (LD) blocks analysis, we finally obtained a total of 79 candidate genes. By detecting the highly associated variations located inside the genic region that overlapped with the results of LD block analysis, we characterized 17 genes that may contribute to salt tolerance during the seed germination stage. At the same time, we conducted a haplotype analysis of the genes with functional variations together with phenotypic correlation and orthologous sequence analyses. Among these genes, <I>OsMADS31</I>, which is a MADS-box family transcription factor, had a down-regulated expression under the salt condition and it was predicted to be involved in the salt tolerance at the rice germination stage. Our study revealed some novel candidate genes and their substantial natural variations in the rice genome at the germination stage. The GWAS in rice at the germination stage would provide important resources for molecular breeding and functional analysis of the salt tolerance during rice germination.</P>

Association of Lipoprotein-Associated Phospholipase A_2 with Characteristics of Vulnerable Coronary Atherosclerotic Plaques

Yu-Sheng Liu,Qing-Hua Lu,Xiao-Bo Hu,Hong-Zhuan Li,Wei-Dong Jiang,Xin Wang,Hao Lin,Ai-Qiong Qin,Yong-Mei Wang,Tong Zhao,Zhao-Qiang Dong,Mei Zhang 연세대학교의과대학 2011 Yonsei medical journal Vol.52 No.6

Purpose: Lipoprotein-associated phospholipase A2 (Lp-PLA_2) is an inflammatory enzyme expressed in atherosclerotic plaques. We investigated the association of circulating Lp-PLA_2 with characteristics of vulnerable coronary atherosclerotic plaques. Materials and Methods: We recruited 113 patients with either unstable angina (UA, n=59) and stable angina (SA, n=54) by coronary angiography. Thirty-six healthy subjects served as controls. Intravascular ultrasound (IVUS) was used to evaluate the characteristics of coronary atherosclerotic plaque, and serum Lp-PLA2 concentration was measured as well. Results: Lp-PLA_2 concentration was significantly higher in both UA and SA patients [(396±36) μg/L and (321±39) μg/L, respectively]compared with the controls [(127±49) μg/L, p<0.01], and higher in UA than SA group. IVUS findings showed that remodeling index (RI) (0.91±0.15 vs. 0.85±0.11, p=0.005) and eccentricity index (EI) (0.73±0.16 vs. 0.65±0.22, p=0.039) were larger in UA than in SA group, and fibrous caps were thicker in SA than UA group [(0.91±0.23) mm vs. (0.63±0.21) mm, p=0.032]. Moreover, Lp-PLA_2 correlated positively with EI (r=0.439, p<0.01) and RI (r=0.592, p<0.05) in UA group. There was an inverse relationship between Lp-PLA_2 and fibrous cap thickness in both UA (r=-0.587, p<0.001) and SA (r=-0.318, p<0.05) groups. The independent risk factors in UA group were Lp-PLA_2 (OR=1.055, 95% CI: 1.03-1.08, p=0.013), LDL-cholesterol (OR=0.032, 95% CI: 0.00-0.05, p=0.041) and fibrous cap thickness (OR=0.008, 95% CI: 0.00-0.45, p=0.019). Lp-PLA_2 was strongly associated with both EI and fibrous cap thickness in both groups. Conclusion: Serum level of Lp-PLA_2 is associated with both eccentricity index and fibrous cap thickness in both UA and SA groups. Elevated levels of circulating Lp-PLA_2 might to be a strong risk factor and more serious for unstable angina than stable angina.

CdS/CdSe lateral heterostructure nanobelts by a two-step physical vapor transport method

Kim, Yu Lee,Jung, Jae Hun,Yoon, Hyun Sik,Song, Man Suk,Bae, Se Hwan,Kim, Yong,Chen, Zhi Gang,Zou, Jin,Joyce, Hannah J,Gao, Qiang,Tan, Hark Hoe,Jagadish, Chennupati IOP Pub 2010 Nanotechnology Vol.21 No.14

<P>The two-dimensional heterostructure nanobelts with a central CdSe region and lateral CdS structures are synthesized by a two-step physical vapor transport method. The large growth rate difference between lateral CdS structures on both ± (0001) sides of the CdSe region is found. The growth anisotropy is discussed in terms of the polar nature of the side ± (0001) surfaces of CdSe. High-resolution transmission electron microscopy reveals the CdSe central region covered with non-uniform CdS layer/islands. From micro-photoluminescence measurements, a systematic blueshift of emission energy from the central CdSe region in accordance with the increase of lateral CdS growth temperature is observed. This result indicates that the intermixing rate in the CdSe region with CdS increases with the increase of lateral CdS growth temperature. In conventional CdSSe ternary nanostructures, morphology and emission wavelength were correlated parameters. However, the morphology and emission wavelength are independently controllable in the CdS/CdSe lateral heterostructure nanobelts. This structure is attractive for applications in visible optoelectronic devices. </P>