采用胚挽救获得Ogura CMS甘蓝与甘蓝型油菜的种间杂种

为将甘蓝型油菜细胞质雄性不育系(Ogura CMS)的恢复基因转至甘蓝Ogura CMS材料上,以6份甘蓝Ogura CMS材料为母本,以含恢复基因的甘蓝型油菜为父本,人工杂交结合胚挽救培养,研究取材时期、培养基成分和杂交组合对胚珠成苗的影响,同时对胚挽救培养获得的植株是否为真实F1杂种进行鉴定。结果发现,胚珠培养以剥蕾授粉后16d取材时胚珠成苗数最多,成苗率为4.56%;培养基以MS+GA 0.1 mg·L^-1+NAA 0.1 mg·L^-1+0.5%水解酪蛋白(CH)+0.5%活性炭(AC)成苗效果最好,成苗率高达6.24%;M09CMS×RFO-46组合成苗数最多,成苗率为5.96%。67株胚挽救培养得到的植株经流式细胞仪、SSR分子标记和形态学鉴定,得出65株为真杂种,真杂种率高达97%。     

基于家族基因分析的甘蓝MLPK互作PUB蛋白的筛选

MLPK(M–位点受体激酶)是芸薹属自交不亲和正向调控关键元件,其参与自交不亲和信号传导的分子机制尚不明确,同时自交不亲和下游信号元件也有待于进一步分离。为了探索分离MLPK互作蛋白的思路和方法,构建了不含核定位信号的MLPK短截蛋白(MLPK-T),并利用酵母双杂交检测到MLPK与臂重复蛋白1(ARC1)作用,通过全基因组鉴定分别获得了96个甘蓝、101个白菜、70个琴叶拟南芥和62个拟南芥PUB蛋白,其中含有臂重复序列的PUB蛋白共为127个。通过系统进化分析,筛选到8个含臂重复序列的甘蓝BoPUB蛋白,其8个基因全部在柱头内表达,且成功利用酵母双杂交检测到MLPK与3个含臂重复序列的BoPUB蛋白Bol008579、Bol016165和Bol023511相互作用。     

亚洲玉米螟核型多角体病毒分离株鉴定及其对草地贪夜蛾的室内毒力测定

本文对从湖北省恩施州亚洲玉米螟幼虫上新分离的一种广谱病毒进行了形态学和分子特征鉴定,并以该病毒与另一种广谱杆状病毒对草地贪夜蛾幼虫进行了生物活性测定。结果表明新分离的核型多角体病毒与薄荷灰夜蛾核型多角体病毒和芹菜夜蛾核型多角体病毒具有较高的同源性,但遗传距离分析不支持它与这两种病毒为同一个种。生物活性测定表明,分离的病毒对草地贪夜蛾3龄幼虫的半数致死剂量是甘蓝夜蛾核型多角体病毒的3.86倍。本研究的结果为开发防治草地贪夜蛾的高效病毒杀虫剂提供了依据。     

红富士苹果芽变系DNA甲基化研究

以35份富士苹果(Malus×domestica Borkh.‘Fuji’)芽变材料为试材,利用甲基化敏感扩增多态性(Methylation Sensitive Amplified Polymorphism,MSAP)分析和UPGMA聚类方法,对其基因组甲基化修饰水平、变异模式以及表观遗传变异关系进行研究。结果表明:(1)不同富士系得到不同的MSAP扩增,总DNA甲基化水平27.90%～36.16%,平均32.87%,双链全甲基化为主要甲基化方式;(2)富士芽变材料绝大多数位点保持了原有甲基化模式;(3)绝大多数芽变(68.57%)检测到全部的甲基化变异模式(12种),去甲基化频率极显著高于甲基化频率(P <0.01),且CG去甲基化极显著高于CHG;(4)36份种质遗传相似系数平均值0.89(0.79～0.92),在聚类图上,富士原种分布在芽变系集中区外,新近发生的芽变系更倾向于聚在一起,着色系片红型和条红型芽变呈分散排布状态。总的来看,富士芽变的甲基化变异模式丰富,超甲基化和去甲基化相伴发生,但以去甲基化为主;‘富士’着色芽变与其最原始品种富士,以及芽变之间发生了较大表观遗传变异;片红和条红型芽变聚类未表现明显偏好性。本研究将为进一步开展富士着色系芽变机理研究提供指导,可以CG去甲基化为切入点展开深入研究。     

有机态和无机态硼对柑橘枳橙砧木生长及生理的影响

以柑橘枳橙[Citrus siinensis(L.)Osb.×Poncirus trifoliata(L.)Raf.]砧木为试验材料,采用水培方式,研究有机态山梨醇—硼和无机态硼酸对其生长及各种生理指标的影响。两种形态硼都能增加砧木幼苗各部位的干、鲜质量和株高;有机态山梨醇—硼对地下部生长的促进作用尤为明显。有机态山梨醇—硼比无机态硼酸更易于向叶片等地上部位转运,有机态山梨醇—硼处理根和茎中的硼含量与无机态硼酸处理没有显著差异,但叶片中的硼含量与硼积累量均显著高于无机态硼酸处理,叶片占总硼含量的比例也显著高于无机态硼酸处理。有机态山梨醇—硼处理各部位钾、钙、镁元素含量高于对照,而无机态硼酸处理各部位元素含量则比有机态山梨醇—硼处理高。缺硼易导致叶片中糖类物质的积累,有机态山梨醇—硼比无机态硼酸能更有效地促进叶片中多糖物质的转运,有机态山梨醇—硼处理可溶性糖含量比对照和无机态硼酸处理分别降低26.3%和11.8%,淀粉含量分别降低43.5%和26.1%。在提高枳橙砧木叶片抗逆能力方面,有机态山梨醇—硼效果优于无机态硼酸,两种硼类型处理叶片中脯氨酸和伤害率均显著低于对照,有机态山梨醇—硼和无机态硼酸分别降低了20.6%和19.6%,有机态山梨醇—硼和无机态硼酸处理伤害率降低了13.3%和16.5%,有机态山梨醇—硼处理叶片中丙二醛含量显著低于无机态硼酸处理19.4%。有机态硼对改善枳橙砧木根系的生长发育的作用效果优于无机态硼酸;能有效降低叶片中碳水化合物的含量,有利于光合作用产物顺利转移;而在减少叶片有害物质的积累、保护质膜方面,有机态与无机态硼有同样的效果。     

黄瓜新品种‘粤丰’

 ‘粤丰’是由早熟抗病的‘豫优4号’为母本,与‘渝亮3号’杂交育成的华北型黄瓜一代杂种。生长势强,主侧蔓结瓜,雌花节率高,回头瓜多。瓜条顺直、美观,长棒形,瓜长38.0 cm,横径3.8 cm,肉厚1.2 cm,单瓜质量约390 g,皮色深绿,光泽度好,刺白,瘤小而密,肉质脆,味微甜,商品性好,耐热,抗病、抗逆性强。适合华南地区春秋季种植。     

利用CRISPR/Ca9技术靶向编辑芥蓝BoaZDS

以芥蓝(Brassicaoleraceavar.alboglabra)为材料,以ζ–胡萝卜素脱氢酶(ζ-Carotene desaturase,ZDS)基因为目标基因,建立其CRISPR/Cas9基因组编辑体系。在BoaZDS的编码区近5′端选择靶位点,构建了CRISPR/Cas9表达载体,通过农杆菌介导的遗传转化方法获得了19个芥蓝转基因阳性植株,Sanger测序分析发现其中13株成功突变,CRISPR/Cas9载体在芥蓝上的突变效率为68.42%,且所有突变植株均表现出明显的白化表型。     

Effects of monocrotaline-induced pulmonary hypertension on expression of Hippo signaling pathway-related molecules in rat lung

AIM:To investigate the expression of Hippo signaling pathway-related molecules in the lung tissues of the rats with pulmonary hypertension induced by monocrotaline for exploring the significance of Hippo signaling pathway in the development of pulmonary hypertension. METHODS:SD rats (n=45) were randomly divided into control group (n=15) and model group (n=30). The rats in model group was given neck subcutaneous injection of monocrota-line at 60 mg/kg to establish pulmonary hypertension model, and the rats in control group was injected with the same volume of normal saline. Four weeks later, right ventricular systolic pressure (RVSP) was measured by right cardiac catheterization, and right ventricular hypertrophy index (RVHI) and right ventricular mass index (RVMI) were calculated. The remodeling of the pulmonary arterioles was observed by HE staining, and medial thickness/external diameter (M/E%) was evaluated. The fibrosis of lung tissues was detected by Masson staining. The protein expression of Yes-associated protein (YAP), tafazzin (TAZ) and TEAD was detected by immunohistochemistry, and the protein and mRNA levels of YAP, TAZ and TEAD in lung tissues were determined by Western blot and RT-qPCR. RESULTS:Compared with control group, the vascular wall in model group was thickened significantly, the M/E% was increased (P<0.01), the pulmonary fibrosis was obvious, and the RVSP and RVHI in model group were significantly higher than those in control group (P<0.01). The immunohistochemical staining showed that the protein expression of YAP, TAZ and TEAD in the pulmonary arterioles in model group was significantly higher than that in control group. The YAP, TAZ and TEAD protein and mRNA levels in the lung tissues were also higher than those in control group (P<0.05). CONCLUSION:The activation of Hippo signaling molecules may promote the remodeling of pulmonary arterioles and further regulate the development of monocrotaline-induced pulmonary hypertension.     

Cepharanthine induces autophagy via PI3K/AKT/mTOR signaling pathway in ovarian cancer SKOV3 cells

AIM: To investigate the autophagy of human ovarian cancer SKOV3 cells induced by cepharanthine and to explore its mechanism. METHODS: The effect of cepharanthine on the viability of ovarian cancer SKOV3 cells was measured by CCK-8 assay. The SKOV3 cells were treated with cepharanthine, and then the formation of autophagosome was observed with acridine orange staining under fluorescence microscope. The protein levels of LC3, AKT, p-AKT, mTOR, p-mTOR and GAPDH in the SKOV3 cells treated with cepharanthine were determined by Western blot.RESULTS: Cepharanthine significantly inhibited the viability of ovarian cancer SKOV3 cells in a dose-dependent manner (P<0.05). The number of the intracellular acidic autophagosomes with bright red fluorescence was significantly increased after cepharanthine treatment in the SKOV3 cells. The expression of LC3-Ⅱ in SKOV3 cells was significantly enhanced after cepharanthine treatment. Furthermore, treatment with cepharanthine in the SKOV3 cells also resulted in a significant down-regulation of phosphorylated form of AKT and mTOR (P<0.01), while the total protein level was not changed. Combination of cepharanthine and 3-methyladenine resulted in a substantial decrease in the cell viability compared with using cepharanthine alone.CONCLUSION: Cepharanthine significantly inhibits the growth of human ovarian cancer SKOV3 cells and induces the autophagy, which may be correlated with down-regulation of PI3K/AKT/mTOR signaling pathway.     

NRF2 attenuates oxidative stress and lysosomal dysfunction in doxorubicin-induced H9C2 cells

AIM:To study the effect of nuclear factor E2-related factor 2 (NRF2) on oxidative stress injury and lysosomal dysfunction in doxorubicin (DOX)-induced rat myocardial H9C2 cells. METHODS:The H9C2 cells were treated with DOX. The expression of NRF2 at mRNA and protein levels was determined by real-time PCR and Western blot. The H9C2 cells stably over-expressing NRF2 were established by lentiviral infection. Real-time PCR and Western blot were used to identify the efficiency of over-expression. After DOX treatment, the cell viability was measured by CCK-8 assay, the activity of lactate dehydrogenase (LDH), superoxide dismutase (SOD), glutathione peroxidase (GSH-Px) and catalase (CAT), and the content of malondialdehyde (MDA) in the cell supernatant were detected. FITC-dextran was used to analyze lysosomal pH, and the protein expression of lysosomal-associated membrane protein 1 (LAMP1) and cathepsin B was determined by Western blot.RESULTS:The expression of NRF2 at mRNA and protein levels in DOX-treated H9C2 cells was significantly decreased (P<0.05). Over-expression of NRF2 significantly up-regulated the mRNA and protein expression of NRF2 in DOX-treated H9C2 cells (P<0.05). After DOX treatment, the cell viability was decreased, and LDH activity was increased. The activity of SOD, GSH-Px and CAT was decreased, and the content of MDA was increased (P<0.05). The lysosomal pH was increased, and the protein expression of LAMP1 and cathepsin B decreased (P<0.05). Over-expression of NRF2 increased the cell viability, decreased LDH activity, increased the activity of SOD, GSH-Px and CAT, and decreased the content of MDA in cell supernatant (P<0.05). Over-expression of NRF2 also decreased the lysosomal pH, and increased the protein expression of LAMP1 and cathepsin B (P<0.05). CONCLUSION:DOX inhibits the expression of NRF2 in the myocardial H9C2 cells. Over-expression of NRF2 attenuates oxidative stress and lysosomal dysfunction in the H9C2 cells induced by DOX.