山羊TNNT3基因可变剪切及其对骨骼肌细胞分化的作用

【目的】快速骨骼肌肌钙蛋白T（fast skeletal troponin T3,TNNT3）作为肌钙蛋白(troponin, Tn)家族成员,调节横纹肌收缩、参与骨骼肌的生长发育并影响家畜肉质性状。通过获得山羊TNNT3基因的可变剪切体,分析山羊TNNT3基因可变剪切的表达模式及其在肌细胞分化中的作用,深入解析TNNT3基因在山羊骨骼肌生长发育过程中的作用机制。【方法】基于NCBI已公布山羊TNNT3基因（NM_001314210.1）和牛TNNT3基因（XM_010821200）mRNA序列,使用软件Primer Premier 6.0设计引物,以简州大耳羊胚胎期和出生后7个阶段骨骼肌为试验材料,克隆测序获得山羊TNNT3基因的CDS区可变剪切体,利用软件ORF Finder、EditSeq、DNAMAN、ClustalW和MEGA_X_10.1.8等对序列进行生物信息学分析;进一步设计实时荧光定量（real-time PCR,RT-qPCR）及半定量引物,研究TNNT3基因剪切体在7个不同组织（背最长肌（longissimus dorsi muscle,LD）、半膜肌（semimembranosus muscle,SM）、心、肝、脾、肺、肾）和7个发育阶段（胚胎期E75、E90、E105和出生后B3、B45、B150、B300）肌肉组织（背最长肌和半膜肌）中表达模式;此外,对转录本TNNT3_3进行体外编码能力检测确定其具有编码蛋白的能力,并在山羊骨骼肌卫星细胞（skeletal muscle satellite cells,MuSCs）中过表达,观察细胞形态变化以及检测标志基因的表达变化,研究其对山羊MuSCs分化的作用。【结果】①TNNT3（NM_001314210.1）CDS区全序列主要含有18个外显子,其中外显子16/17相互排斥,转录后单一表达。克隆发现山羊TNNT3基因 5个新转录本（TNNT3_1—5）,其外显子数分别是15、15、20、16、14。②生物信息学分析结果显示山羊TNNT3基因核苷酸序列和氨基酸序列与绵羊、牛、猪等哺乳动物具有很高的一致性,而与鱼类和爬行类动物的一致性较低,说明TNNT3基因序列在哺乳动物高度保守。③TNNT3 mRNA在背最长肌、半膜肌、心、肝、脾、肺、肾7个组织中都有表达,其中在骨骼肌中高度富集(P < 0.01),心脏及肺次之,其余组织中较低;TNNT3 mRNA在背最长肌和半膜肌中的表达始终处于一个动态变化中,胚胎期TNNT3在半膜肌的表达量高于背最长肌（P<0.05）;出生后则背最长肌中高于半膜肌（P<0.05）。④山羊TNNT3基因转录本TNNT3_3重复出现保守的外显子9—11（138bp）,体外翻译实验显示其可编码蛋白且蛋白大小与预期基本相符（37 kD）;相较于对照组,在山羊MuSCs中过表达该转录本使肌分化标志基因Myomaker、MyoG和MyH4 mRNA极显著升高(P < 0.01)。【结论】获得了山羊TNNT3基因具有完整CDS区5个新可变剪切体,TNNT3主要在肌肉组织（背最长肌和半膜肌）中高表达,在哺乳动物中高度保守且促进成肌分化。初步表明TNNT3基因在动物肌肉生长发育中具有重要的生物学功能。     

Effects of different lighting on reproductive system in depressive female rats

AIM: To investigate the effects of different lighting on the reproductive system in depressive female rats. METHODS: Healthy adult female rats were randomly chosen as control group, and the depressive adult female rats in SPF grade were randomly divided into 5 groups(7 rats each):depressive model group, sulfur lamp group, heat radiation lamp group, fluorescent lamp group and LED lamp group. After 45 d of continuous illumination, the estrous cycle was observed by the vaginal exfoliated cells, and the organ indexes of ovary and uterus were calculated. The concentrations of estiadrol(E₂), prolactin(PRL), progesterone(PROG) and testosterone(T) in the serum were detected by ELISA, and the histopathological lesion of ovary was observed under microscope with HE staining. RESULTS: The estrous cycle exhibited serious disorder, the ovaries exhibited serious congestion, and the organ indexes of ovary and uterus and the concentrations of E₂, PRL, PROG and T decreased significantly in the rats in depressive model group compared with control group(P<0.05). The estrous cycle and histopathological damage of ovary were obviously improved, and the concentrations of E₂, PRL, PROG and T were significantly increased after the sulfur lamp lighting in the depressive female rats compared with depressive model group. No obvious change and improvement of the reproductive functions in the heat radiation lamp group, fluorescent lamp group and LED lamp group was observed. CONCLUSION: The reproductive functions of depressive female rats are improved by sulfur lamp lighting.     

Habitat associations of saplings and adults in an old-growth temperate forest in the Changbai mountains,northeastern China

Species-habitat association analysis is useful to detect spatial arrangement of individual plants, to discover rules about the distribution of species and to generate hypotheses about the possible underlying process controlling observed structures. Quantifying methods were used to classify habitats in terms of topographical variables in a mixed temperate broad-leaved Korean pine forest of the Changbai mountains in northeastern China. All of the 625 20 m × 20 m quadrats of the plot could be unambiguously assigned to one of three habitat categories (low-plateau, high-plateau and slope). Torus-translation tests were used to estimate species-habitat associations. Many species are clearly distributed in a biased fashion with respect to habitats. Fifteen (55.6%) out of 27 species showed strong positive or negative association with specific habitats. We compared species-habitat associations at the sapling and adult stages. Adjusted density values indicated few species exhibit extremely strong habitat associations. Only 9 out of 26 species had adjusted densities > 3 in the habitat for which they had strong positive affinity. Few species show the same associations at the small tree and large tree stages. Only 3 out of 22 occurring associations with a specific habitat appeared to have a consistent habitat association at the two stages. These results suggest that species-habitat associations exist in the 25-ha plot of the temperate forest of the Chang- bai mountains. Owing to limitations in our statistical methodology, we partly underestimated associations by ignoring rare species. Regeneration niches can contribute to co-existence, but regeneration niches due to habitat associations play a limited role in species co-existence, since most species show a similar trend in habitat associations at the sapling and adult stages. We should pay more attention to shifts in habitat associations, i.e. niche shifts at different stages of existence.     

中金系列杨树的组培技术研究

中金系列杨树是杨柳科杨属白杨派植物,选择其腋芽（或顶芽）、嫩茎段、嫩叶片等为外植体,进行愈伤组织诱导试验、不定芽分化试验、生根试验,筛选出了适合植株分化、生根的培养基成分;对生根试管苗进行移栽基质配比试验,确定组培苗最佳生长环境。总结出适合中金系列杨树组培的技术流程,保证了杨树的遗传稳定性,降低了培养成本,加快了培养速度。     

Seasonal patterns of photochemical capacity and spring phenology reveal genetic differentiation among native Scots pine (Pinus sylvestris L.) populations in Scotland

Environment-driven genetic differentiation among populations is a common feature among forest trees, and an understanding of how populations have adapted to their home site conditions is essential for management and conservation practices. In Scotland, 84 native Scots pine (Pinus sylvestris L.) woodlands are recognised by the Forestry Commission and they occupy highly diverse environments from the maritime west coast to continental sites in eastern Scotland. However, it is not known whether adaptations to local environments along sharp temperature and rainfall gradients have occurred in different populations and as a result, the seed transfer guidelines of the species are based only on data from isozymes and monoterpenes. In this study of an outdoor common-garden trial, we used chlorophyll fluorescence to examine whether seedlings from 32 open-pollinated families and eight populations from sites experiencing contrasting annual temperature regimes differed in their response to variation in natural outdoor temperatures between September 2009 and May 2010. In addition, growth initiation in spring was recorded. Photochemical capacity at photosystem II F_v/F_m showed a distinct seasonal trend and remained at relatively high levels (∼0.7) until November. Following a period of over 2 weeks with temperatures below or close to 0 °C, F_v/F_m started decreasing towards its minimum values recorded in early March when population means varied between 0.35 and 0.45. By early May and along with rising temperatures, photochemical capacity had recovered to the same level as observed in early November. Populations were found to respond differently to the cold period starting in December. The largest drop in photochemical capacity was observed in seedlings from a low-altitude population located in the maritime western Scotland, while in seedlings from higher-altitude locations in the cooler eastern Scotland, the response was smaller. In March, the recovery of photochemical capacity was slowest in seedlings from the mildest and coolest sites. Evidence of adaptive genetic differentiation was also found in spring phenology. Initiation of shoot elongation and needle flush were earlier in families from higher altitudes (cooler areas), but population differences were not significant at the α = 0.05 level. These results suggest that adaptation to the spatially heterogeneous environment in Scotland has taken place in Scots pine and that in order to minimise the risk of planting maladapted seed stock, the patterns of environmental and adaptive genetic variation should be taken into account in the management of genetic resources in this species.     

Effects of astragaloside on the levels of sex hormone and oxidative stress injury in rats with polycystic ovary syndrome

AIM To investigate the effects of astragaloside on the levels of sex hormone and oxidative stress in rats with polycystic ovary syndrome （PCOS）. METHODS Female SD rats （n=60） were randomly divided into normal control group, model group, Diane-35 （0.339 2 mg/kg） group, low dose astragaloside （12.5 mg/kg） group and high dose astragaloside （50 mg/kg） group, with 12 rats in each group. The PCOS model was induced by letrozole （1 mg/kg）, which was administered by gavage once a day for 3 weeks. After administration, the estrus cycle of the rats was observed by vaginal smear, and the ovarian index was calculated. HE staining was used to observe the histopathological changes of the ovaries. Serum levels of the sex hormones testosterone （T）, estradiol （E₂）, luteinizing hormone （LH） and follicle-stimulating hormone （FSH） were measured by ELISA. The levels of superoxide dismutase （SOD）, malondialdehyde （MDA） and glutathione peroxidase （GSH-Px） in serum and ovarian tissue were detected by colorimetry, and the protein levels of steroidogenetic acute regulatory protein （StAR） and apoptosis-related proteins cleaved caspase-3, Bax and Bcl-2 in ovarian tissue were detected by Western blot. RESULT Compared with control group, the oestrous cycle of the rats in model group was disorder, and the ovarian index was increased, ovary was polycystic. The serum levels of T, LH and MDAwere significantly increased （P<0.05）, while the contents of E2, FSH and the activities of GSH-Px and SOD were significantly decreased （P<0.05）. The levels of MDA, StAR, cleaved caspase-3 and Bax proteins in ovarian tissue were significantly up-regulated （P<0.05）. GSH-Px and SOD activities and Bcl-2 protein levels were significantly down-regulated （P<0.05）. CONCLUSION Astragalosideeffectively balances the levels of sex hormone in PCOS rats and relieves the oxidative stress injury, the mechanism may be related to the inhibition of StAR expression.     

玉米灰斑病菌致病性分化研究

从我国玉米品种资源中首次成功筛选出9个具有广泛血缘代表性的自交系,即沈137,78599-1,Mo17,478,C8605-2,E28,598,Va35,K12,作为玉米灰斑病菌的生理分化鉴别寄主;8个主栽品种,即沈试29,沈试31,铁单9,掖单13,丹413,丹玉18,沈农87,东单54,作为辅助鉴别寄主,从而确定了玉米灰斑病菌生理分化鉴别寄主体系。采用田间成株期鉴定,高粱粒灌心法进行接种,根据在鉴别寄主上的发病等级,将采自北方玉米主产区的23个玉米灰斑病菌菌株划分成5个致病类型。研究表明,我国玉米灰斑病菌存在一定程度的致病性分化,通过病级评价可将23个玉米灰斑病菌菌株分成5个致病类型,其中致病类型I为强致病类型,致病类型IV为弱致病类型,Ⅱ,Ⅲ2组致病类型介于两者之间,而致病类型V属于不确定类型。研究结果为我国玉米品种抗性鉴定、灰斑病流行监测和品种合理布局提供了理论依据。     

高山低温诱导蝴蝶兰花芽分化过程中的生理变化

连续观察，测定了蝴蝶兰花芽分化过程中不同低温天数时植株体内某些生理生化变化，结果显示：低温处理20d后，叶片内矿质元素、可溶性糖、淀粉、可溶性蛋白、游离氨基酸、过氧化物酶含量变化显著，说明低温诱导过程中叶片内进行着一系列生理生化变化，为花芽分化提供物质和能量。     

Indica-japonica differentiation in Chinese rice landraces

Summary Ninety Chinese rice landraces were examined with special reference to the indica-japonica differentiation in terms of traditional criteria, isozyme analysis and PCR analysis of the chloroplast DNA (cpDNA). Cultivars were separated into indica and japonica defined by a discriminant function (Z) based on key characters, as well as by isozyme genotypes. Most indica landraces had chloroplast DNAs with a deletion at the Pst-12 fragment, while most japonica landraces had cpDNAs without the deletion. Two traditionally recognized varietal groups in China, keng and hsien, corresponded largely to the respective japonica and indica revealed in our study. The results obtained in this study showed good agreement for classification of indica and japonica types by the three methods: discriminant analysis by Z value, isozyme analysis, and PCR analysis for cpDNA.