银杏锰型超氧化物歧化酶GbMnSOD基因的克隆与表达

利用RACE技术首次从银杏中克隆到锰型超氧化物歧化酶基因(GbMnSOD ) 的cDNA全长。GbMnSOD的cDNA全长965 bp (GenBank accession number: EF633506) 。生物信息学分析GbMnSOD cDNA序列含有一个681 bp最大读码框, 编码一个226氨基酸多肽链, 通过软件预测分子量为2515 kD, 等电点为8197。三维结构预测结果显示, GbMnSOD 含有12个α螺旋和3 个β折叠构成一个篮子状的活性中心。GbMnSOD氨基酸序列与其它植物MnSOD具有很高的相似性。进化树分析结果表明GbMnSOD和其他物种的MnSOD源自于相同的祖先。Southern杂交显示, GbMnSOD属于一个小的多基因家族。Northern 杂交表明GbMnSOD在银杏的根、茎、叶和果中都有表达, 在叶中的表达量最高, GbMnSOD 的转录受到ABA、IAA、蔗糖、甘露醇、NaCl 和低温的诱导。     

ERF转录因子新成员JERF3提高百合的耐盐性

以麝香百合鳞片愈伤组织为受体，通过对农杆菌浓度、培养时间及抗生素敏感性的比较试验，建立了农杆菌介导的麝香百合愈伤组织遗传转化的优化体系，并利用该体系将ERF转录因子JERF3基因导入百合中，通过PCR、RT-PCR及Southern blot检测，证明JERF3已整合到百合基因组中并在转录水平上表达。对T0代转基因株系耐盐性鉴定表明，与未经转化的对照相比，转基因百合的耐盐性能明显提高，说明JERF3基因的超表达能有效提高转基因百合抵抗盐害的能力。     

白菜β-1,3 - 葡聚糖酶基因cDNA的克隆及序列分析

 以白菜抗霜霉病品种‘雪克青’cDNA 为模板, 采用RT2PCR 技术, 获得了1032 bp 的β- 1,3- 葡聚糖酶基因的cDNA 序列。序列分析表明, 克隆的β- 1,3 - 葡聚糖酶基因cDNA 序列编码343 个氨基酸, 与芜菁bg1 基因具有98 %的同源性, 与拟南芥bg2 基因具有84%的同源性。在GenBank 中登录号为AY395720 。     

新高系梨10个品种S基因型的鉴定

以亲缘关系极近的新高系梨10个品种及被视为‘新高’亲本的‘天之川’和‘今村秋’梨等为试验材料，对其基因组DNA进行了PCR-RFLP检测、DNA序列测定及生物信息学分析。结果表明：新高系梨10个品种即‘新高’、‘黄金’、‘水晶’、‘早生黄金’、‘早蜜新高’、‘天皇’、‘金秋’、‘晚大新高’、‘农家新高’和‘鲜黄’的S基因型分别为S₃S₉、S₃S₄、S₃S₉、S₃S₄、S₃S₉、S₃S₉、S₃S₉、S₃S₉、S₃S₉和S₃S₅；‘天之川’、‘今村秋’的S基因型分别为S₁S₉和S₁S₆，通过基因型判定‘今村秋’非‘新高’梨的父本。     

甘蓝型油菜DOF蛋白基因的克隆与生物信息学分析

Dof转录因子是植物特有的一类转录因子,含有一个独特的单锌指结构域,在植物的生长发育中起重要的调控作用。以甘蓝型油菜为试材,采用电子克隆和RT-PCR技术,以期克隆一个具有Zf-Dof结构域的蛋白基因cDNA序列。结果表明:该基因含有1284bp的开放阅读框,编码427个氨基酸,蛋白分子量为46.9kDa,等电点为8.86,为亲水性蛋白。经序列比对分析表明其与拟南芥CDF3基因有较高同源性,可能具有相似的功能。     

甜瓜果实酸性转化酶基因cDNA片段的克隆

 根据在GenBank中登录的番茄、胡萝卜和柑桔等酸性转化酶基因的保守序列设计引物，采用RT-PCR方法，从甜瓜果实总RNA中扩增出目标cDNA片段，克隆到pMD18-T载体中。序列分析表明，它与其它植物的酸性转化酶基因的同源性很高，与番茄氨基酸序列同源性为99％，说明已经成功克隆到甜瓜果实酸性转化酶基因cDNA片段，在GenBank中登记号AF490425。     

葡萄MADS-box家族基因保守片段的克隆与序列分析

MADS-box转录因子在多种植物的发育过程特别是花器官和果实发育过程中发挥重要的作用。为了从葡萄中克隆出新的MADS-box基因,研究在葡萄花及果实发育中的作用,根据多种植物MADS-box基因保守区序列,设计简并引物,应用RT-PCR技术从红富士葡萄花序中分离出34条MADS-box基因cDNA片段。序列分析表明,这些片段长度在138～152bp之间,包含基因起始密码子。推导的氨基酸序列与已登录的欧洲葡萄及其它物种的MADS-box基因同源性超过83%。用推导的氨基酸序列与已知的欧洲葡萄和拟南芥MADS-box家族基因进行系统发育分析,可将这些基因片段分别归入拟南芥MADS-box基因不同亚家族中,证明葡萄中存在多种MADS-box家族基因,克隆的片段包含ABCDE模型中的各类花发育基因。     

白菜GLDH 基因cDNA的克隆及序列分析

 以白菜品种‘苏州青’ cDNA 为模板, 采用RT-PCR、巢式PCR、3’RACE和5’RACE技术, 获得了L - 半乳糖酸- 1, 4 - 内酯脱氢酶(EC 1.3.2.3, GLDH) 基因cDNA 2 034 bp全长序列。序列分析表明, GLDH基因cDNA序列编码601个氨基酸。其氨基酸序列与花椰菜GLDH基因具有98%的同源性, 与拟南芥GLDH基因具有90%的同源性。该基因在GenBank中登录号为AY899298。     

马蔺DFR基因的克隆及生物信息学特征分析

以马蔺花瓣为试材,通过转录组测序,应用PCR技术克隆马蔺花青素生物合成关键酶二羟黄酮醇-4-还原酶(dihydvroflavonol-4-reductase,DFR)基因并获得其生物信息学特征。结果表明:马蔺DFR基因全长1 427bp,共编码357个氨基酸(该基因命名为IlDFR,GenBank登录号为KY907171)。该基因在氨基酸水平上与多种植物的DFR具有较高的同源性,在系统进化上与同属的荷兰鸢尾(Iris hollandica)的DFR亲缘关系较近;预测其蛋白质相对分子质量为39.99kDa,等电点为5.89,主要由α-螺旋和β-折叠构成,定位于细胞质,属于酸性亲水不含信号肽的不稳定类蛋白质。马蔺IlDFR有一段氨基酸序列‘VTGASGYVGSWLVMKLLRDGY’与大部分物种的DFR特有相对保守的NADPH结合域非常相似,只有4个氨基酸残基有所差异。     

芜菁花叶病毒外壳蛋白基因的克隆与序列分析

将来自北京地区甘蓝和芥菜上的TuMV外壳蛋白基因片段分别克隆到pMD18-T载体中,测序并进行序列分析。结果表明:两者CP基因序列长度均为867个核苷酸,编码288个氨基酸,它们的核苷酸同源性为97·2%,氨基酸同源性为97.6%;两者与GenBank中的部分CP基因序列的核苷酸同源性在87.8%~97.8%之间,氨基酸同源性在91.7%~99.0%之间;分子进化树分析两者可归属World-B组。     

Effects of radiation from 188Re on smooth muscle cell proliferation

AIM: Although endovascular radiotherapy inhibits neointimal hyperplasia, the exact alterations induced by β-particles irradiation remain to be elucidated. The objective of this study was to investigate the ability and the cellular mechanism of local β^-particles emission from ¹⁸⁸Re to inhibit vascular smooth muscle cells (SMCs). METHODS: The SMCs in vitro were irradiated by ¹⁸⁸Re with single doses of 2.6 Gy-25.8 Gy. The effects of β-particles on SMCs, such as effective irradiate doses, the period of inhibition for SMCs proliferation, the changes of cell proliferation rate and DNA synthesis rate, cell cycle progression and related gene expression, were investigated by cell count, [³H]-TdR incorporation, cell cycle progression analysis, cell viability and immunocytochemistry, respectivecy. RESULTS: β-particles irradiation with dose of 5.2 Gy could inhibit significantly SMCs proliferation. At dose of 20.6 Gy DNA synthesis inhibitory rate was 92%, SMCs proliferation rate was only 3%. Renoval of ¹⁸⁸Re did not abolish the inhibitory effects of β-particles on SMCs proliferation. The expression of P53 was up regulation and PCNA was down regulation after irradiation. CONCLUSION: β-particles from ¹⁸⁸ Re was significantly effective and permanent in inhibiting SMCs proliferation, and inhibitory effect was in dose-dependet manner ED50was 5 Gy, the best dose to inhibit SMCs proliferation was 20 Gy. β-particles irradiation induced SMCs to occur G₀/G₁ arrest, damaged the ability of SMCs reproliferation and led to cell clonogenic death. P53 and PCNA had regulatiory effects on SMCs proliferation after β-particles irradiation.     

Effect of L-Arg on plasma content of endothelin and expression of c-fos mRNA in the left ventricle of rats with renovascular hypertensive hypertrophy

AIM:To study the effect of L-Arg on plasma content of endothelin (ET) and the expression of proto-oncogene c-fos mRNA in the left ventricle of rats with renovascular hypertensive hypertrophy. METHODS: The level of c-fos mRNA were measured by in situ hybridization. The ET in plasma were measured by radioimmunoassay. RESULTS:After eight weeks of treatment with L-Arg, the expression of c-fos decreased markedly (P＜0.01). The ET content in plasma also decreased significantly by L-Arg(P＜0.01).CONCLUSION: Plasma ET content and the expression of c-fos in the left ventricle of rats with renovascular hypertensive hypertrophy could be decreased by L-Arg administration.     

Compositional and Functional Properties of Saskatoon Berry and Blueberry

Abstract

Saskatoon berry (Amelanchier alnifolia Nutt., Rosaceae) and blueberry (Vaccinium corymbosum L., Ericaceae) are substantially equivalent in all characteristics that are important to the consumer, including fruit color, shape, size, nutrition, texture, and uses. In addition, both fruits are native to North America and they have practically identical historical uses and known health benefits. Their composition, processing, nutritional value and metabolism, intended uses, and levels of undesirable substances are compared.     

Cryopreservation of shoot apices of hawthorn in vitro cultures originating from East Asia

The objective of this study was to establish a cryopreservation protocol for hawthorn shoot apices (Crataegus pinnatifida Bge.). Cryopreservation was carried out via encapsulation–dehydration, vitrification, and encapsulation–vitrification on shoot apices excised from in vitro cultures. We began by showing that cold-acclimation enhanced the regrowth of cryopreserved apices from 10.0 to 65.5% in encapsulation–dehydration. We then decided that the encapsulation–dehydration method was an optimal cryopreservation method for hawthorn shoot apices in terms of its high recovery after cryopreservation as well as its ease of use compared with vitrification and encapsulation–vitrification. In encapsulation–dehydration, the protocol leading to optimal regrowth was as follows: after cold-acclimation at 5 °C in the dark for 2 weeks, excised shoot tips were pretreated for 24 h at 25 °C on hormone-free Murashige and Skoog [Murashige, T., Skoog, F., 1962. A revised medium for rapid growth and bioassays with tobacco tissue culture. Physiol. Plant. 15, 473–497] (MS) basal medium with 0.4 mol/L sucrose, then encapsulated and precultured in liquid MS medium with 0.8 mol/L sucrose for 16 h at 25 °C. Precultured beads were dehydrated for 6 h at 25 °C in the dessicator containing 50 g silica gel to a moisture content of 15.3% (fresh-weight basis) before cryostorage for 1 h. In addition, we examined the effect of adding glycerol to both the alginate beads and loading solution to enhance regrowth after cryopreservation in encapsulation–dehydration. In the present study, it was shown that adding 0.5 mol/L glycerol resulted in high regrowth percentages (82.5–90.0%) in four Crataegus species.     

多效唑对猕猴桃离体试管苗生长及内源激素的影响

多效唑（ＰＰ３３３）处理猕猴桃试管苗，降低了其生长强度；植株体内的ＧＡ３、ＩＡＡ和ＺＴ含量下降，ＡＢＡ的含量上升，乙烯释放率增加；并且能降低外源的ＧＡ３和ＩＡＡ促进生长的作用，而外源的ＧＡ３和ＩＡＡ又能不同程度地逆转多效唑的抑制作用，使植株恢复生长。     

Proteomic analysis between pathological scars and normal skin

AIM: To investigate and screen the sensitive proteins in the formation mechanism of pathological scars by comparing the results of differential proteomic analysis between pathological scars and normal skin.METHODS: Two-dimensional gel electrophoresis was used to detect the protein expression profiles in 8 keloid patients, 8 hypertrophic scar patients and 3 matched normal skin patients.The proteins that showed differential expression of over 4-fold change were cut and analyzed by MALDI-TOF/TOF mass spectrometry.RESULTS: A two-dimensional protein profiling comparison between pathological scars and normal skin was successfully established.On average, 2 978 spots in keloid, 2 975 spots in hypertrophic scar and 3 053 spots in normal skin were identified using gel analysis software.Compared with normal skin, there were totally 36 differentially-expressed proteins in keloid and hypertrophic scar identified from the spots of over 4-fold change, including 16 proteins in both keloid and hypertrophic scar (8 up-regulated and 8 down-regulated), 11 only in keloid (9 up-regulated and 2 down-regulated) and 9 only in hypertrophic scar (4 up-regulated and 5 down-regulated).CONCLUSION: Proteomic analysis can identify the proteins with variance of pathological scars versus normal skin, thus providing probable new clues to reveal the formation mechanism of pathological scars.     

Metallothionein inhibits homocysteine-induced proliferation of rat vascular smooth muscle cells

AIM:To investigate the effect of metallothionein(MT) on proliferation of rat vascular smooth muscle cells (VSMCs) stimulated by homocysteine and its mechanism. METHODS:VSMCs proliferation was measured by [³-H]-TdR incorporation, mitogen-activated protein kinase(MAPK)activity were determined by immunoprecipitation method, the intracellular contents of MT and malondialdehyde (MDA)were assayed by -hemoglobin saturation method and TBA reaction, respectively, and lactate dehydrogenase (LDH) leakage was measured by NADH oxidation. RESULTS:Hcy(10^-6-10^-4 mmol/L) stimulated [³-H]-TdR incorporation by the VSMCs in a concentration-dependent manner. Compared with control, [³-H]-TdR incorporation in VSMCs treated with 0.1 mmol/L Hcy was increased by 4.2 fold (P＜0.01). Meanwhile, Hcy enhanced MAPK activity, MDA formation and LDH release (P＜0.01)in a concentration-dependent manner. Treatment of VSMCs with MT alone did not change above parameters, compared with control. However, MT (10^-6-10^-4 mol/L)attenuated significantly Hcy-stimulated proliferation of VSMCs (P＜0.01)in a concentration-dependent manner. And MT inhibited obviously Hcy-induced activation of MAPK activity, MDA formation and LDH release. Preincubation of VSMCs with 0.5 mmol/L ZnCl₂ for 6 h induced an increase cellular MT content by 5.7-fold (P＜0.01). The MT-overexpressed VSMCs resisted Hcy-stimulating action on MAPK activity, MDA formation and LDH leakage (P＜0.01). CONCLUSION:These results show that MT has an inhibitory effect on Hcy-induced VSMCs proliferation, and that MT could inhibit Hcy-stimulated MAPK activity and lipid peroxidation.     

Coupling past management practice and historic landscape change on John F. Kennedy Space Center,Florida

Historic landcover dynamics in a scrubby flatwoods (Tel-4) and scrub landscape (Happy Creek) on John F. Kennedy Space Center were measured using aerial images from 1943, 1951, 1958, 1969, 1979, and 1989. Landcover categories were mapped, digitized, geometrically registered, and overlaid in ARC/INFO. Both study sites have been influenced by various land use histories, including periods of range management, fire suppression, and fire management. Several analyses were performed to help understand the effects of past land management on the amount and spatial distribution of landcover within the study sites. A chi-squared analysis showed a significant difference between the frequency of landcover occurrence and management period. Markov chain models were used to project observed changes over a 100-year period; these showed current management practices being effective at Tel-4 (restoring historic landscape structure) and much less effective at Happy Creek. Documenting impacts of past management regimes on landcover has provided important insight into current landscape composition and will provide the basis for improving land management on Kennedy Space Center and elsewhere.     

XBP-01 accelerated apoptosis induced by oxidized low density lipoprotein in vascular smooth muscle cells

AIM: Previous studies performed with XBP-01 in vitro indicated that XBP-01 could inhibit vascular smooth muscle cells from being transformed into foam cell and could eliminate the atherosclerotic plaque in C57BL/6J mouse. This experiment is to investigate its mechanism of eliminating plaques in vitro. METHODS: The cultured porcine artery smooth muscle cells incubated with XBP-01 of 0.1 mg/L for 24 h after preincubated with oxidized low density lipoprotein of 15 mg/L for 72 h in vitro. The samples were analyzed by fluorescence microscope, confocal microscope system and flow cytometry. RESULTS: Apoptosis was triggered by being incubated with oxidized low density lipoprotein and this process was accelerated additionally by being incubated with XBP-01. CONCLUSION: XBP-01 can be effective in eliminating atherosclerotic plaque by accelerating the process in which oxidized low density lipoprotein induced smooth muscle cell apoptosis.     

Influence of Sini decoction on the proliferation and apoptosis of artery smooth muscle cells after balloon injury

AIM: To investigate the influence of Sini decoction (SND) on the proliferation and apoptosis of rabbit abdominal aorta smooth muscle cells after ballon injury and discuss the effect of vascular smooth muscle cell's (VSMCs) proliferation and apoptosis in post-percutaneous coronary intervention (PCI) restenosis (RS) and the feasibility of SND preventing post-PCI RS. METHODS: The animal model of rabbit abdominal aorta ballon injury was set up and the therapertic group was treated with SND. The shape of proliferative and apoptotic cell were investigated by electron microscope. Immunohistochemistry staining was performed using α-actin,PCNA and Cyclin E monoclonal antibodies. In situ Cell Death Detection Kit was used to identify apoptotic cells. Abdomial aorta angiography was operated in the 84th day subgroup and the stenosis degree was evalued by quantitative angiographic analysis. RESULTS: As compared with the control group, the therapeutic group displayed a lower proliferative percentage and a higher apoptosic percentage (P<0.05). Moreover, the apoptosic peek time was on the 14th day after operation,which was longer than the control group. CONCLUSION: SND effectly inhibited the proliferation of VSMCs and iuduced apoptosis in VSMCs.