Effect of antisense oligonucleotides targeting telomerase catalytic subunit on tumor cell proliferationin vitro

To screen specific antitumor drugs targeting telomerase catalytic subunit (hEST2), 12 different hEST2 antisense oligonucleotides were designed based on hEST2 mRNA second structure and transfected into tumor cell lines by the lipofectin-mediated method. Cell growth activity was evaluated by MTT assay. hEST212 was picked out and its specificity, antitumor tree and continuous effect were analyzed. The results showed that hEST212 had promising antitumor activity in vitro, hEST2 can be used as a pratical target and an antisense drug candidate for cancer.     

Cloning and functional research of renal cell carcinoma related novel gene—— GYLZ-RCC18

After the renal cell carcinoma related novel gene fragmentGYLZ-RCC18 was cloned by using suppression subtractive hybridization (SSH), we used the SMART RACE technology to clone the full length ofGYLZ-RCC18 and performed chromosome location by the FISH method. RT-PCR was used to detect the expression of the first reading frame ofGYLZ-RCC18 in different stages and grades of renal cell carcinoma tissue and other tissues. Also we transfected the antisense oligonucleotide ofGYLZ-RCC18 to renal cell carcinoma cell line GRC-1, and analyzed the proliferation activity, growth speed, apoptosis and mortality changes in GRC-1. The results show that the full length ofGYLZ-RCC18 (GenBank accession No.: BE825133) cDNA is about 3.5 kb long which is located at No. 14 chromosome.GYLZ-RCC18 has a higher expression in higher grades and stages of renal cell carcinoma than in the lower ones. The expression ofGYLZ-RCC18 in renal cell carcinoma was much higher than that in normal kidney and other tissues. After transfection ofGYLZ-RCC18 antisense oligonucleotide, the mortality of GRC-1 increases evidently, the proliferation activity and growth speed were inhibited remarkably at the same time. Also the antisense oligonucleotide can induce the apoptosis of GRC-1 all through the observation time. Our results indicated thatGYLZ-RCC18 is an important novel gene related to renal cell carcinoma. Its overexpression would stimulate the growth and proliferation activity and plays an antidead and antiapoptosis effect in renal cell carcinoma. Transfection of antisense oligonucleotide could inhibit the generation and development of renal cell carcinoma. The study provides a new clue for the research of renal cell carcinoma, and also provides an instruction for special genetic diagnosis and the therapy of renal cell carcinoma.     

北部湾广西海域海洋真菌多样性及其生物活性研究

本研究从北部湾广西海域采集的海绵、珊瑚、沉积物等样品中分离真菌,旨在挖掘北部湾的海洋真菌资源,筛选出具有潜在抗肿瘤或抗菌活性的菌株。采用稀释涂布法和基于内转录间隔区基因序列(ITS)的系统发育树法分析海洋真菌的多样性信息,并通过滤纸片琼脂扩散法和MTT比色法评价菌株的抗菌、抗肿瘤活性。分离得到45株海洋真菌,隶属17属。其中青霉属(Penicillium)和曲霉属(Aspergillus)为优势种群,分别占总菌株种类的26.7%和24.4%。此外,获得一株潜在的新种菌株Myrothecium gramineum GXIMD01018。活性筛选结果发现菌株Aspergillus japonicus GXIMD01014、Penicillium oxalicum GXIMD01021、Talaromyces purpureogenus GXIMD01024等6株真菌的次级代谢产物对人结直肠癌细胞具有较显著的细胞毒活性,菌株T.purpureogenus GXIMD01024的代谢粗提物具有一定的抗耐甲氧西林金黄色葡萄球菌(Methicillin-Resistant Staphylococc...     

S 100A8 mediates the activation of P65/HLA- B/S 100A8/BCL-2/Caspase-9 （-3） pathway in laryngeal carcinogenesis

S100 calcium binding protein A8 （S100A8）, a possible novel member of NF-kappa B signal pathway in laryngeal squamous cell carcinoma （LSCC）, interacts with human leukocyte antigen B （HLA-B） which carries an NF-kappa B binding site within the enhancer A. The objective of this study was to explore the molecular mechanism of S100A8 in laryngeal carcinogenesis. RT-PCR, Western blotting and immunohistochemistry staining were applied to evaluate the expression levels of IKKα, P65, REL-B, S100A8, APAF-1 and BCL-2 genes. The signal transduction passway in which S100A8 might participate was explored by RNA interference. Flow cytometry, TUNEL assay and cell invasion in vitro were used to detect the biological behavior of Hep2 cells induced by S100A8 gene. Our results showed that high expression of S100A8 was related to tumorigenesis in LSCC and negatively correlated with the degree of differentiation, indicating that S100A8 gene could inhibit apoptosis and promote metastasis in LSCC. Additionally, the suppression of S100A8 by RNA interference down-regulated BCL-2 but not APAF-1, P65 and IKKα, while, the suppression of P65 could significantly down-regulate the expression of S100A8 gene. In conclusion, S100A8 plays an important role in P65/HLA-B/S100A8/BCL-2/Caspase-9 （-3） pathway in laryngeal carcinoma.     

3种苏铁属植物总黄酮、总多糖含量及抗氧化活性研究

为探究锈毛苏铁(Cycas ferruginea)、叉孢苏铁(C.segmentifida)和石山苏铁(C.sexseminifera) 3种苏铁属(Cycas)植物的总黄酮、总多糖含量以及抗氧化活性,本研究采用超声波辅助法对其根、叶柄、叶、雄球花、茎干的总黄酮及总多糖含量进行提取,以DPPH自由基(DPPH·)、羟自由基(·OH)、超氧阴离子自由基(O^-₂·)清除率评价其抗氧化活性,并采用Pearson相关性分析法分析相关性。结果表明：锈毛苏铁、叉孢苏铁和石山苏铁叶的总黄酮含量较高,大小依次为锈毛苏铁(8.61 mg·100 mg^-1)>叉孢苏铁(7.82 mg·100 mg^-1)>石山苏铁(1.04 mg·100 mg^-1);茎干中总多糖含量最高,大小依次为叉孢苏铁(28.32 mg·100 mg^-1)>石山苏铁(24.43 mg·100 mg^-1)>锈毛苏铁(16.59 mg·100 mg^-1<...     

Reporter-based screen for <Emphasis Type="Italic">Arabidopsis</Emphasis> mutants compromised in nonhost resistance

Plants are exposed to many potentially pathogenic microbes in the environment, but each species is only susceptible to a limited number of pathogens. The broad resistance is referred to as nonhost resistance. To date, little is known about the underlying mechanism of nonhost resistance and the signaling transduction process. Here we describe a simple method for isolating Arabidopsis nonhost resistance mutants against a nonadapted bacterial pathogen. A RAP2.6 promoter-driven LUC reporter system was developed to replace the tedious bacterial growth assay during the primary screening. The RAP2.6-LUC reporter gene is normally induced by the virulent bacterium Pseudomonas syringae pv tomato but not the nonadapted bacterium P. syringae pv phaseolicola. By using this method we iso- lated 4 mutants displaying strong reporter activity in response to P. syringae pv phaseolicola, which were characterized in some details, ebsl, ebs2, ebs3, and ebs4 （enhanced bacterial susceptibility） were compromised in resistance against P. syringae pv phaseolicola and/or P. syringae pv tomato. In addition, ebs4 showed enhanced hypersensitive response to the incompatible bacterium P. syringae pv tomato （avrB）. These results demonstrated that the method is suited for large scale screening for nonhost resistance mutants.     

邻香草醛缩胱氨酸双Schiff碱的合成及其对粟酒裂殖酵母细胞生长代谢的热动力学研究

用摩尔比为2:1的邻香草醛(C_8H_8O_3)与L-胱氨酸(C_6H_(12)N_2O_4S_2)反应,合成了一种新的双Schiff碱化合物--双{2-[(3-巯基丙酸钠)-2-亚胺基-甲基]-6-甲氧基-苯酚}(OVCS)。通过元素分析、红外光谱、核磁共振等手段对其组成和结构进行了表征,确定其化学式为Na_2(C_(22)H_(22)N_2O_8S_2),采用TAM air微量热仪测定了新合成的Schiff碱化合物(OVCS)在305.15 K时对粟酒裂殖酵母细胞作用的产热曲线;根据产热曲线计算了在OVCS作用下,粟酒裂殖酵母细胞生长代谢的最大发热功率p_(max)、速率常数k、传代时间tG、抑制率I和半抑制浓度C_(I,50)等热动力学参数。通过实验可以发现随着OVCS浓度的增加,粟酒裂殖酵母细胞的生长代谢速率常数k、生长代谢的总热效应Q_(total)、最大发热功率p_(max)均减小,抑制率I、达到生长代谢最大功率所需时间t_(max)、传代时间tG均增加等规律,半抑制浓度C_(I,50)为35.99 mg/L(或9.62×10~(-2)mol/L)。实验结果表明,OVCS对粟酒裂殖酵母细胞有抑制作用,且浓度越大,抑制作用越强。     

转录因子OsNAC2对逆境下水稻产量性状的影响

以水稻OsNAC2过表达、RNAi转基因株系和野生型（日本晴）为材料,分别在苗期和生殖期进行干旱和盐胁迫处理,探索逆境条件下OsNAC2对水稻产量性状的影响。结果表明,不论是在苗期还是生殖期,OsNAC2-RNAi株系的叶相对含水量均比野生型更高,对干旱胁迫具有更强的适应能力;而OsNAC2过表达株系则相反。虽然苗期和生殖期遭遇盐胁迫的OsNAC2-RNAi株系相比野生型具有更高的叶相对含水量,但是OsNAC2的过量表达与RNAi株系的产量性状跟野生型相比并没有明显不同。生殖期干旱和盐胁迫下转基因株系的产量性状分析显示：干旱胁迫下,OsNAC2-RNAi株系的结实率与野生型相比显著提高了20.8%~29.2%,千粒重则无明显差异;而OsNAC2过表达株系每株粒数和千粒重相比野生型株系均显著降低。虽然盐胁迫下OsNAC2-RNAi株系的分蘖数和有效穗数明显比野生型高,但单株粒数和千粒重则无明显差异。上述结果表明,OsNAC2-RNAi株系具有更强的耐旱性,对于干旱胁迫下水稻的产量有显著的提高作用。     

乳苣水提取物对人肺癌细胞SPCA-1生长的影响

为了探讨乳苣水提取物对体外培养的肺癌细胞SPCA-1的影响,分别用质量浓度为0.5、1、1.25和1.5 g/L(mg/m L)的乳苣水提取物处理SPCA-1细胞,24 h后通过CCK-8法检测发现,与对照组(未处理)相比,处理组细胞增殖活力均显著下降(P0.001).流式细胞仪检测(AnnexinⅤ-FITC/PI染色)发现随着处理浓度的增加,处理组细胞凋亡率都显著提高(P0.05),且呈剂量依赖关系.为了探索乳苣是否对体内肿瘤有抑制作用,构建了SPCA-1细胞的裸鼠皮下瘤模型,通过观察喂食乳苣水提取物后皮下瘤的生长情况发现,与未喂食乳苣水提取物的对照鼠相比,喂食5周后的实验组的瘤体得到明显的抑制.实验结果表明乳苣水提取物能够抑制体外和体内生长的SPCA-1细胞,可以作为一种潜在的预防和治疗肺癌的药物做进一步研究.     

条斑紫菜与坛紫菜挥发性风味的电子鼻分析

【目的】利用电子鼻技术区分条斑紫菜(Pyropia yezoensis)与坛紫菜(Pyropia haitanensis)的原藻及其制品。【方法】应用18个金属氧化型气体传感器并通过风味雷达图和主成分分析法对2种紫菜的挥发性风味进行轮廓性分析。【结果】两种方法分析原藻及其制品均显示条斑紫菜和坛紫菜的气味轮廓与敏感物质类型存在差异;相比紫菜原藻,各制品间的气味轮廓较为一致。【结论】电子鼻技术可以区分条斑紫菜与坛紫菜挥发性风味,原藻及其制品均有明显差异。加工过程使得产品挥发性风味趋于统一。     

Fine mapping of the red plant gene R1 in upland cotton(Gossypium hirsutum)

Sub 16 is a substitution line with G. hirsutum cv. TM-1 genetic background except that the 16th chromosome (Chr. 16) is replaced by the corresponding homozygous chromosome of G. barbadense cv. 3-79, and T586 is a G. hirsutum multiple gene marker line with 8 dominant mutation genes. The R ₁ gene for anthocyanin pigmentation was tagged in Chr. 16 in T586. The objective of this research was to screen SSR markers tightly linked with R ₁ by using the F₂ segregating population containing 1259 plants derived from the cross of Sub 16 and T586 and the backbone genetic linkage map from G. hirsutum×G. barbadense BC₁ newly updated by our laboratory. Genetic analysis suggested that the segregation ratio of red plants in the F₂ population fit Mendelian 1:2:1 inheritance, confirming that the red plant trait was controlled by an incomplete dominance gene. Preliminary mapping of R ₁ was conducted using 237 randomLy selected F₂ individuals and JoinMap v3.0 software. Then, a fine map of R1 was constructed using the F₂ segregating population containing 1259 plants, and R ₁ was located between NAU4956 and NAU6752, with only 0.49 cM to the nearest maker loci (NAU6752). These results provided a foundation for map-based cloning of R ₁ and further development of cotton cultivars with red fibers by transgenic technology. Supported by National Natural Science Foundation of China (Grant No. 30730067) and Programme of Introducing Talents of Discipline to Universities (Grant No. B08025)     

产丁醇大肠杆菌工程菌的构建

【目的】为了在大肠杆菌(Escherichia coli)中导入改良的丁醇合成途径,使非生产菌株大肠杆菌具备产丁醇的能力。【方法】克隆大肠杆菌乙酰转移酶基因atoB和丙酮丁醇梭菌(Clostridium acetobutylicum)丁醇合成途径关键酶基因(crt、hbd、adhE),构建多顺反子表达质粒pSE380-atoB-adhE-crt-hbd;克隆齿垢密螺旋体(Treponema denticola)反式烯酰辅酶A还原酶基因ter,构建表达质粒pSTV29-ter,并将双质粒导入到大肠杆菌。【结果】构建的工程菌能半厌氧发酵产微量丁醇,产量为0.08g/L。【结论】大肠杆菌中的丁醇合成途径导入成功,构建了产丁醇的大肠杆菌工程菌。     

拟南芥雄性不育突变体opw(only pollen wall)候选突变基因的克隆

在T-DNA插入突变体Salk_059463株系的群体中,筛选到两株雄性不育突变体,对TDNA序列上的一对引物进行PCR鉴定,结果表明:其基因组中没有T-DNA插入.遗传分析表明这两株雄性不育突变体由同一单个隐性基因控制,引起不育的主要原因是从花药发育的第8期开始,小孢子细胞质内容物逐渐减少直至消失,到花药发育的第12期,药室内的小孢子只剩下一个花粉壁空壳,故该突变体命名为opw(only pollen wall).利用图位克隆的方法对OPW基因进行了定位,结果表明OPW基因位于第二条染色体上分子标记T28M21和T3G21之间的12 kb区间内,该区间内一共有21个基因注释.通过克隆区间内的基因并测序发现opw-1突变体基因组中At2g40140基因编码序列的外显子在第289和第290个碱基之间插入了一个A碱基,而opw-2突变体基因组中At2g40140基因编码序列的外显子在第412和第413个碱基之间插入了一个T碱基,造成的编码序列移码使第424至第426碱基成为终止密码子,故At2g40140是编码OPW的候选基因.     

南亚热带杉木、红锥人工林碳储量及分配特征

明确南亚热带杉木(Cunnighamia lanceolata)、红锥(Castanopsis hystrix)人工林碳储量及分配特征,可为应对全球气候变化研究提供基础数据,为碳汇林业发展提供科学依据。以我国亚热带地区广泛栽培的杉木人工林和红锥人工林为研究对象,以相对生长方程计算林木生物量,实测林下植被生物量、林木和林下植被各组分含碳率、土壤含碳率等,进而分析不同人工林的碳储量及分配规律。结果表明:(1)人工林生态系统不同组分的含碳率存在一定差异,虽然杉木和红锥的全株含碳率相差无几,分别为48.04%和47.80%,但林下植被和土壤表层的含碳率差别较大,林下植被含碳率为40.84%—47.73%(杉木林)、36.69%—43.76%(红锥林);土壤表层含碳率为2.28%—3.30%;(2)杉木人工林乔木层碳储量(71.48t/hm~2)、林下植被碳储量(1.533t/hm~2)显著高于红锥人工林乔木层碳储量(51.82t/hm~2)和林下植被碳储量(1.185t/hm2),而红锥人工林枯落物层碳储量(0.673t/hm2)显著高于杉木人工林(0.386t/hm~2);(3)杉木人工林的皮、叶、根碳储量显著高于红锥人工林,相反,红锥人工林的枝碳储量(8.04t/hm~2)显著高于杉木人工林(6.00t/hm~2);(4)杉木人工林生态系统碳储量(217.56t/hm~2)与红锥人工林生态系统碳储量(195.05t/hm~2)无显著差异,土壤和乔木层是人工林生态系统的主要碳库,分别占生态系统碳储量的66.37%—72.81%和26.59%—32.93%。杉木人工林乔木层、林下植被和生态系统碳储量均高于红锥人工林,红锥人工林枯落物碳储量显著高于杉木人工林,杉木是发展碳汇林的较好树种。     

优良豆科牧草大翼豆06-2选育研究

以豆科牧草大翼豆[Macroptilium atropur-pureum(dc.)urbcv.Siratro]为材料,通过单株选择,培育出优质高产的豆科牧草大翼豆06-2[M.atropur-pureum(dc.)urbcv.Siratro No.06-2],并对其进行品种比较试验和区域试验。结果表明,大翼豆06-2适应性广,抗寒,耐轻霜,丰产性能好,年平均鲜草产量高达42822.0kg/hm2,比对照品种高20.3%~33.3%。此外,大翼豆06-2生长速度快,蛋白质含量高达19.0%,具有较高的营养价值,有望成为亚热带地区的优良豆科牧草品种。     

Ubi1 intron-mediated enhancement of the expression of Bt cry1Ah gene in transgenic maize （Zea mays L.）

The cry1Ah gene was one of novel insecticidal genes cloned from Bacillus thuringiensis isolate BT8. Two plant expression vectors containing cry1Ah gene were constructed. The first intron of maize ubiqutinl gene was inserted between the maize Ubiquitin promoter and cry1Ah gene in one of the plant expressing vectors （pUUOAH）. The two vectors were introduced into maize immature embryonic calli by microprojectile bombardment, and the reproductively plants were acquired. PCR and Southern blot analysis showed that foreign genes had been integrated into maize genome and inherited to the next generation stably. The ELISA assay to T1 and T2 generation plants showed that the expression of CrylAh protein in the construct containing the ubil intron （pUUOAH） was 20% higher than that of the intronless construct （pUOAH）. Bioassay results showed that the transgenic maize harboring cry1Ah gene had high resistance to the Asian corn borers and the insecticidal activity of the transgenic maize containing the ubil intron was higher than that of the intronless construct. These results indicated that the maize ubil intron can enhance the expression of the Bt cry1Ah gene in transgenic maize efficiently