Selection and evaluation of reference genes for qRT-PCR analysis of different developmental stages in Chilo suppressalis

He Fu-Jing,Lu Ming-Xing,Du Yu-Zhou 한국응용곤충학회 2021 Journal of Asia-Pacific Entomology Vol.24 No.4

Quantitative real-time RT-PCR is considered to valuable method for the analysis of gene expression. But for the data accuracy of gene expression, the normalization of data using reference genes is essential. In this study, we focus on the analysis of the relative expression of nine commonly used candidate reference genes in the different developmental stages and female and male pupae/adults of Chilo suppressalis, and expression stabilities were analyzed by the ΔCt method, geNorm, NormFinder, and RefFinder. The results showed that H3 was the most stable reference genes in the different developmental stages while EF1 was the optimal genes in pupae/adult of different sexes of C. suppressalis. To validate, the expression profile of the gene encoding heat shock protein 60 (Hsp60) was investigated. The results showed that the transcription of target gene Hsp60 varied depending on the reference gene selected for normalization in different developmental stages, but the trend of expression of Hsp60 between the most stable reference gene combination and the least stable reference gene in pupae/adults of different sexes was the same. Therefore, it is imperative to select the reference genes to estimate target gene expression accurately in the specific experiment. This study provides scientific basis on functional analyses of the important genes contributed to the development and sex differentiation of C. suppressalis.

A Novel Xylanase from Streptomyces sp. FA1: Purification, Characterization, Identification, and Heterologous Expression

Jie He,Lingqia Su,Xiaojun Sun,Jiajia Fu,Jian Chen,Jing Wu 한국생물공학회 2014 Biotechnology and Bioprocess Engineering Vol.19 No.1

A xylanase (XynA) was purified from the culturemedium of Streptomyces sp. FA1, which was previouslyisolated from a bamboo retting system. XynA had amolecular mass of 43 kDa, displayed maximal activity atpH 5.5, retained 41% of its maximal activity at pH 11.0,and was stable over a wide pH range (3.0 ~ 11.0). PurifiedXynA was subjected to peptide mass fingerprinting, whichled to the cloning of the xynA gene. The xynA gene, whichencodes a mature protein of 436 amino acids, washeterologously expressed in E. coli BL21(DE3). The activityin the culture medium could reach 213.5 U/mL, which was11.2-fold higher than that produced by Streptomyces sp. FA1. BLAST searching revealed that full-length XynAshares less than 90% identity with most of its homologues,whereas amino acids 48-436 of the enzyme share 97%identity with an open reading frame encoding a putativefull-length mature xylanase from Streptomyces tendae. Thetruncated xynA gene, xynA48-436, was cloned and expressedin E. coli, however, no xylanase activity could be detectedin the culture medium. Based on these results, it is suggestedthat XynA is a new member of glycoside hydrolasesfamily10 with exceptional catalytic efficiency at alkalinepH.

Complete Mitochondrial Genome of a Tongue Worm Armillifer agkistrodontis

Jian Li,Fu-Nan He,Hong-Xiang Zheng,Rui-Xiang Zhang,Yi-Jing Ren,Wei Hu 대한기생충학열대의학회 2016 The Korean Journal of Parasitology Vol.54 No.6

Armillifer agkistrodontis (Ichthyostraca: Pantastomida) is a parasitic pathogen, only reported in China, which can cause a zoonotic disease, pentastomiasis. A complete mitochondrial (mt) genome was 16,521 bp comprising 13 protein-coding genes (PCGs), 22 tRNA genes, 2 rRNA genes, and 1 non-coding region (NCR). A phylogenetic tree drawn with the concatenated amino acid sequences of the 6 conserved PCGs (atp6, cox1-3, and nad2) showed that A. agkistrodontis and Armillifer armillatus constituted a clade Pentastomida which was a sister group of the Branchiura. The complete mt genome sequence of A. agkistrodontis provides important genetic markers for both phylogenetic and epidemiological studies of pentastomids.

Ca<SUP>2⁢</SUP>-induced Ca<SUP>2⁢</SUP> Release from Sarcoplasmic Reticulum Negatively Regulates Myocytic ANP Release in Beating Rabbit Atria

Dan Li,He Xiu Quan,Jin Fu Wen,Jing Yu Jin,Sung Hun Park,Sun Young Kim,Sung Zoo Kim,Kyung Woo Cho 대한생리학회-대한약리학회 2005 The Korean Journal of Physiology & Pharmacology Vol.9 No.2

It is not clear whether Ca<SUP>2⁢</SUP>-induced Ca<SUP>2⁢</SUP> release from the sarcoplasmic reticulum (SR) is involved in the regulation of atrial natriuretic peptide (ANP) release. Previously, we have shown that nifedipine increased ANP release, indicating that Ca<SUP>2⁢</SUP> entry via voltage-gated L-type Ca<SUP>2⁢</SUP> channel activation decreases ANP release. The purpose of the present study was two-fold: to define the role of SR Ca<SUP>2⁢</SUP> release in the regulation of ANP release and whether Ca<SUP>2⁢</SUP> entry via L-type Ca<SUP>2⁢</SUP> channel is prerequisite for the SR-related effect on ANP release. Experiments were performed in perfused beating rabbit atria. Ryanodine, an inhibitor of SR Ca<SUP>2⁢</SUP> release, increased atrial myocytic ANP release (8.69⁑3.05, 19.55⁑1.09, 27.31⁑3.51, and 18.91⁑4.76% for 1, 2, 3, and 6μM ryanodine, respectively; all P<0.01) with concomitant decrease in atrial stroke volume and pulse pressure in a dose-dependent manner. In the presence of thapsigargin, an inhibitor of SR Ca<SUP>2⁢</SUP> pump, ryanodine-induced increase in ANP release was not observed. Thapsigargin attenuated ryanodine-induced decrease in atrial dynamic changes. Blockade of L-type Ca<SUP>2⁢</SUP> channel with nifedipine abolished ryanodine-induced increase in ANP release (0.69⁑5.58% vs. 27.31⁑3.51%; P<0.001). In the presence of thapsigargin and ryanodine, nifedipine increased ANP release and decreased atrial dynamics. These data suggest that Ca<SUP>2⁢</SUP>-induced Ca<SUP>2⁢</SUP> release from the SR is inversely involved in the regulation of atrial myocytic ANP release.

Endothelial Aquaporin-1 (AQP1) Expression Is Regulated by Transcription Factor Mef2c

Yong Jiang,He Liu,Wen-jing Liu,Hai-bin Tong,Chang-jun Chen,Fu-gui Lin,Yan-hang Zhuo,Xiao-zhen Qian,Zeng-bin Wang,Yu Wang,Peng Zhang,Hong-liang Jia 한국분자세포생물학회 2016 Molecules and cells Vol.39 No.4

Aquaporin 1 (AQP1) is expressed in most microvascula-ture endothelial cells and forms water channels that play major roles in a variety of physiologic processes. This study aimed to delineate the transcriptional regulation of AQP1 by Mef2c in endothelial cells. Mef2c cooperated with Sp1 to activate human AQP1 transcription by binding to its proximal promoter in human umbilical cord vein endothelial cells (HUVEC). Over-expression of Mef2c, Sp1, or Mef2c/Sp1 increased HUVEC migration and tube-forming ability, which can be abolished AQP1 knockdown. These data indicate that AQP1 is a direct target of Mef2c in regulating angiogenesis and vasculogenesis of endothelial cells.

Ca2+-induced Ca2+ Release from Sarcoplasmic Reticulum Negatively Regulates Myocytic ANP Release in Beating Rabbit Atria

Dan Li,He Xiu Quan,Jin Fu Wen,Jing Yu Jin,,박성훈,김선영,김성주,조경우 대한약리학회 2005 The Korean Journal of Physiology & Pharmacology Vol.9 No.2

It is not clear whether Ca2+-induced Ca2+ release from the sarcoplasmic reticulum (SR) is involved in the regulation of atrial natriuretic peptide (ANP) release. Previously, we have shown that nifedipine increased ANP release, indicating that Ca2+ entry via voltage-gated L-type Ca2+ channel activation decreases ANP release. The purpose of the present study was two-fold: to define the role of SR Ca2+ release in the regulation of ANP release and whether Ca2+ entry via L-type Ca2+ channel is prerequisite for the SR-related effect on ANP release. Experiments were performed in perfused beating rabbit atria. Ryanodine, an inhibitor of SR Ca2+ release, increased atrial myocytic ANP release (8.69±3.05, 19.55±1.09, 27.31±3.51, and 18.91±4.76% for 1, 2, 3, and 6μM ryanodine, respectively; all P<0.01) with concomitant decrease in atrial stroke volume and pulse pressure in a dose-dependent manner. In the presence of thapsigargin, an inhibitor of SR Ca2+ pump, ryanodine-induced increase in ANP release was not observed. Thapsigargin attenuated ryanodine-induced decrease in atrial dynamic changes. Blockade of L-type Ca2+ channel with nifedipine abolished ryanodine-induced increase in ANP release (0.69±5.58% vs. 27.31±3.51%; P<0.001). In the presence of thapsigargin and ryanodine, nifedipine increased ANP release and decreased atrial dynamics. These data suggest that Ca2+-induced Ca2+ release from the SR is inversely involved in the regulation of atrial myocytic ANP release.

Endothelial Aquaporin-1 (AQP1) Expression Is Regulated by Transcription Factor Mef2c

Jiang, Yong,Liu, He,Liu, Wen-jing,Tong, Hai-bin,Chen, Chang-jun,Lin, Fu-gui,Zhuo, Yan-hang,Qian, Xiao-zhen,Wang, Zeng-bin,Wang, Yu,Zhang, Peng,Jia, Hong-liang Korean Society for Molecular and Cellular Biology 2016 Molecules and cells Vol.39 No.4

Aquaporin 1 (AQP1) is expressed in most microvasculature endothelial cells and forms water channels that play major roles in a variety of physiologic processes. This study aimed to delineate the transcriptional regulation of AQP1 by Mef2c in endothelial cells. Mef2c cooperated with Sp1 to activate human AQP1 transcription by binding to its proximal promoter in human umbilical cord vein endothelial cells (HUVEC). Over-expression of Mef2c, Sp1, or Mef2c/Sp1 increased HUVEC migration and tube-forming ability, which can be abolished AQP1 knockdown. These data indicate that AQP1 is a direct target of Mef2c in regulating angiogenesis and vasculogenesis of endothelial cells.

Association of TNF-α-308 and -238 Polymorphisms with Risk of Cervical Cancer: A Meta-analysis

Pan, Feng,Tian, Jing,Ji, Chu-Shu,He, Yi-Fu,Han, Xing-Hua,Wang, Yong,Du, Jian-Ping,Jiang, Feng-Shou,Zhang, Ying,Pan, Yue-Yin,Hu, Bing Asian Pacific Journal of Cancer Prevention 2012 Asian Pacific journal of cancer prevention Vol.13 No.11

Published data on the associations between tumor necrosis factor-alpha (TNF-${\alpha}$) promoter -308G>A and -238G>A polymorphisms and cervical cancer risk are inconclusive. To derive a more precise estimation of the relationship, a meta-analysis was performed. Data were collected from MEDLINE and PubMed databases. Crude odds ratios (ORs) with 95% confidence intervals (CIs) were calculated in a fixed/random effect model. 13 separate studies including 3294 cases and 3468 controls were involved in the meta-analysis. We found no association between TNF-${\alpha}$-308G>A polymorphism and cervical cancer in overall population. In subgroup analysis, significantly elevated risks were found in Caucasian population (A vs. G: OR = 1.43, 95% CI = 1.00-2.03; AA vs. GG: OR = 2.09, 95% CI = 1.34-3.25; Recessive model: OR = 2.09, 95% CI = 1.35-3.25) and African population (GA vs. GG: OR = 1.53, 95% CI = 1.02-2.30). An association of TNF-${\alpha}$-238G>A polymorphism with cervical cancer was found (A vs. G: OR = 0.61, 95% CI = 0.47-0.78; GA vs. GG: OR = 0.59, 95% CI = 0.45-0.77; Dominant model: OR = 0.59, 95% CI = 0.46-0.77). When stratified by ethnicity, similar association was observed in Caucasian population (A vs. G: OR = 0.62, 95% CI = 0.46-0.84; GA vs. GG: OR = 0.59, 95% CI = 0.43-0.82; Dominant model: OR = 0.60, 95% CI = 0.44-0.83). In summary, this meta-analysis suggests that TNF-${\alpha}$-238A allele significantly decreased the cervical cancer risk, and the TNF-${\alpha}$-308G>A polymorphism is associated with the susceptibility to cervical cancer in Caucasian and African population.

Improvement of the Thermostability of Xylanase from Thermobacillus composti through Site-Directed Mutagenesis

( Yong-sheng Tian ),( Jing Xu ),( Lei-chen ),( Xiao-yan Fu ),( Ri-he Peng ),( Quan-hong Yao ) 한국미생물생명공학회(구 한국산업미생물학회) 2017 Journal of microbiology and biotechnology Vol.27 No.10

Thermostability is an important property of xylanase because high temperature is required for its applications, such as wood pulp bleaching, baking, and animal feedstuff processing. In this study, XynB from Thermobacillus composti, a moderately thermophilic gram-negative bacterium, was modified via site-directed mutagenesis (based on its 3D structure) to obtain thermostable xylanase, and the properties of this enzyme were analyzed. Results revealed that the half-life of xylanase at 65°C increased from 10 to 50 min after a disulfide bridge was introduced between the α-helix and its adjacent β-sheet at S98 and N145. Further mutation at the side of A153E named XynB-CE in the C-terminal of this α-helix enhanced the half-life of xylanase for 60 min at 65°C. Therefore, the mutant may be utilized for industrial applications.

[$Ca^{2+}-induced$ $Ca^{2+}$ Release from Sarcoplasmic Reticulum Negatively Regulates Myocytic ANP Release in Beating Rabbit Atria

Li, Dan,Quan, He Xiu,Wen, Jin-Fu,Jin, Jing-Yu,Park, Sung-Hun,Kim, Sun-Young,Kim, Sung-Zoo,Cho, Kyung-Woo The Korean Society of Pharmacology 2005 The Korean Journal of Physiology & Pharmacology Vol.9 No.2

It is not clear whether $Ca^{2+}-induced$ $Ca^{2+}$ release from the sarcoplasmic reticulum (SR) is involved in the regulation of atrial natriuretic peptide (ANP) release. Previously, we have shown that nifedipine increased ANP release, indicating that $Ca^{2+}$ entry via voltage-gated L-type $Ca^{2+}$ channel activation decreases ANP release. The purpose of the present study was two-fold: to define the role of SR $Ca^{2+}$ release in the regulation of ANP release and whether $Ca^{2+}$ entry via L-type $Ca^{2+}$ channel is prerequisite for the SR-related effect on ANP release. Experiments were performed in perfused beating rabbit atria. Ryanodine, an inhibitor of SR $Ca^{2+}$ release, increased atrial myocytic ANP release ($8.69{\pm}3.05$, $19.55{\pm}1.09$, $27.31{\pm}3.51$, and $18.91{\pm}4.76$% for 1, 2, 3, and $6{\mu}M$ ryanodine, respectively; all P<0.01) with concomitant decrease in atrial stroke volume and pulse pressure in a dose-dependent manner. In the presence of thapsigargin, an inhibitor of SR $Ca^{2+}$ pump, ryanodine-induced increase in ANP release was not observed. Thapsigargin attenuated ryanodine-induced decrease in atrial dynamic changes. Blockade of L-type $Ca^{2+}$ channel with nifedipine abolished ryanodine-induced increase in ANP release ($0.69{\pm}5.58$% vs. $27.31{\pm}3.51$%; P<0.001). In the presence of thapsigargin and ryanodine, nifedipine increased ANP release and decreased atrial dynamics. These data suggest that $Ca^{2+}$-induced $Ca^{2+}$ release from the SR is inversely involved in the regulation of atrial myocytic ANP release.