Characteristics of the mitochondrial genome of Rana omeimontis and related species in Ranidae: Gene rearrangements and phylogenetic relationships

The Omei wood frog (Rana omeimontis), endemic to central China, belongs to the family Ranidae. In this study, we achieved detail knowledge about the mitogenome of the species. The length of the genome is 20,120 bp, including 13 protein‐coding genes (PCGs), 22 tRNA genes, two rRNA genes, and a noncoding control region. Similar to other amphibians, we found that only nine genes (ND6 and eight tRNA genes) are encoded on the light strand (L) and other genes on the heavy strand (H). Totally, The base composition of the mitochondrial genome included 27.29% A, 28.85% T, 28.87% C, and 15.00% G, respectively. The control regions among the Rana species were found to exhibit rich genetic variability and A + T content. R. omeimontis was clustered together with R. chaochiaoensis in phylogenetic tree. Compared to R. amurensis and R. kunyuensi, it was more closely related to R. chaochiaoensis, and a new way of gene rearrangement (ND6‐trnE‐Cytb‐D‐loop‐trnL2 (CUN)‐ND5‐D‐loop) was also found in the mitogenome of R. amurensis and R. kunyuensi. Our results about the mitochondrial genome of R. omeimontis will contribute to the future studies on phylogenetic relationship and the taxonomic status of Rana and related Ranidae species.     

Cloning,expression and characterization of the putative nuclear transport factor 2 (NTF2) gene from moss <Emphasis Type="Italic">Conocephalum conicum</Emphasis>(L.) Dum

Biomacromolecules import into the nucleus is a complex progress which requires the participation of several cytosolic factors, and nuclear transport factor 2 (NTF2) is one of essential components in nuclear trafficking. Its main role is to transport RanGDP from cytoplasm to nucleus by interacting with FxFG nucleoporin repeats. In the study a putative new gene, designated as CcNTF2, was obtained from the moss (Conocephalum conicum) cDNA library we have constructed. The full-length cDNA sequence is 913 bp in size contains a 372 bp open reading frame (ORF) flanked by a 195 bp 5′-untranslated sequence and a long 346 bp 3′-non-coding region, encoding 123 amino acids of 13,575.3 Da. Part of the genomic sequence was also cloned and sequenced, which is 1,602 bp long and possesses two exons and one intron. Alignment analysis showed that the CcNTF2 protein is high conserved among plant NTF2 and shares 81% similarity with the ones from Arabidopsis thaliana and Brassica rapa. The expression of wild-type CcNTF2 was detected by immunoblotting of extraction of C. conicum and it indicated the putative protein is integral. Through functional expression of CcNTF2-green fluorescent protein (GFP) in tobacco, it was demonstrated that CcNTF2 can accumulate at the nuclear rim. Site-directed mutagenesis analysis confirmed CcNTF2 P71K has influence on the protein import into nucleus. In addition, overexpression of CcNTF2 P71K was observed to be deleterious for the plant cell. It is the first illumination of NTF2 in moss, and our study established the primary foundation for further research on moss NTF2.     

柔嫩艾美耳球虫新基因的生物信息学分析与克隆表达

为了研究柔嫩艾美耳球虫(Eimeria tenella)的两个主要入侵发育阶段——子孢子和裂殖子的差异基因,根据已报道的子孢子与裂殖子差异的ESTs序列,应用RACE技术克隆获得了子孢子阶段差异表达的新基因全长cDNA序列,命名为ZL583.该基因全长为862 bp,开放阅读框(ORF)为486 bp,编码161个氨基酸,编码蛋白的分子量约为16.9 kD,利用生物信息学分析软件,分析新基因所编码蛋白的性质、定位、结构等特征.利用荧光定量PCR对柔嫩艾美耳球虫不同发育阶段该基因的表达量进行分析显示,子孢子阶段的表达高于其他发育阶段.将ZL583克隆于pET-28a中构建重组质粒,转化大肠杆菌BL21(DE3)后经IPTG诱导表达,获得重组蛋白分子量约23 kD.免疫印迹分析显示该重组蛋白可与兔抗子孢子血清发生特异性反应,表明该蛋白具有较好的反应原性.该结果为进一步研究该基因的生物学功能奠定基础.     

Agkihpin,a Distinct SVTLE from the Venom of Gloydius halys Pallas: Purification,Characterization and Structure–Activity Determination

Blood clots produced by snake‐venom thrombin‐like enzymes (SVTLEs) are cleared rapidly, which makes SVTLEs attractive as potential candidates for antithrombotic therapy. We isolated a SVTLE, agkihpin, from the venom of Gloydius halys Pallas . Agkihpin was confirmed to a single‐chain TLE with molecular mass of 25.5 kD, pI of 7.43, optimal pH of 8.0 (hydrolyzing TAME), linked carbohydrate absent, and weak fibrinogen clotting activity. It was also found that (i) G. halys might be the latest species in SVTLEs phylogenetic tree; (ii) different level of conservation was shown among the SVTLEs from the Viperidae snakes. Some of those site may account for different activities exhibited by those SVTLEs, especially position 181, at which a fibrinogenolytic activity increase was found when a basic and larger amino acid substituted by a neutral and smaller one; (iii) an extra α‐helix constructed with a ‘Pro + acidic amino acid + aromatic amino acid’ pattern was found in the SVTLEs from Gloydius and Agkistrodon snakes, although it does not necessarily imply an effect on the fibrinogenolytic activity of the SVTLEs. This study provided some new insight into the activity of SVTLE.     

周期应变对动脉平滑肌细胞分泌血管紧张素II的影响

应用自行设计的硅胶膜伸张加载装置对培养于硅胶膜上的Ｗｉｓｔａｒ 大鼠主动脉平滑肌细胞施以周期性二维伸张应变（ 最大伸长比：纵向０ ．７ ％ ,横向０ ．３ ％ ） ,运用放射免疫沉淀法对其ＡｇＩＩ 的分泌量进行测定, 结果表明： 加载组血管紧张素ＩＩ 分泌量明显高于对照组（Ｐ＜ ０ ．００１ ,２ｈ ,１０ｈ 除外） ,对照组分泌曲线较平坦,而加载组曲线较尖锐;加载４ｈ ,ＡｇＩＩ分泌量达到峰值（２１９ ±２０ ．０ｐｇ／１０５ｃｅｌｌｓ） 。结合形态观察,表明力学信号能对主动脉平滑肌细胞的行为产生明显影响     

组胺H1和H2受体在兔灌流肺低氧性肺动脉加压反应中的作用

本实验采用兔自身血液灌流原位肺,用含氧6％的氮氧混合气进行人工通气,在此基础上,分别用扑尔敏和西咪替丁阻滞肺血管的H_1和H_2受体,观察组胺受体在低氧性肺动脉加压反应(HPPR)中的作用,并测定肺流出血液中血浆cAMP和cGMP含量。结果表明,扑尔敏可使兔灌流肺HPPR减弱,西咪替丁则使之增强。提示组胺参与兔灌流肺的HPPR;H_1和H_2受体在兔的HPPR中以H_2受体的作用占优势;血浆环核苷酸含量变化,可能是兔低氧时肺动脉收缩及组胺发生影响的生化基础之一。