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基于Xcm I酶切的pUC19-T载体的构建 总被引：1，自引：0，他引：1
[目的]构建以pUC19质粒为基础可利用Xcm I内切酶制备的T栽体。[方法]化学合成2条含有双XcmI酶切位点的互补寡聚核苷酸链,经过变性、复性后克隆入pUCl9质粒的HindIII和BamHI位点之间,通过XcmI酶切后得到一个线性化的带有3’末端突出一个T碱基的T栽体。[结果]经TA克隆验证,制备的pUC19-HB-T载体对PCR产物的克隆率达到95％以上。[结论]构建的pUC19-HB-T载体可用于PCR产物的克隆、测序及后续分子生物学操作。 相似文献
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AIM: To investigate the feasibility to inhibit the expression of MHCⅡ by special siRNA targeting class Ⅱ major histocompatibility complex (MHC Ⅱ) transactivator (CⅡTA), which might regulate MHC Ⅱexpression for suppressing immune rejection. METHODS: Five different siRNA were designed, synthesized and transfected into freshly isolated rat corneal keratocytes. At 24 hours posttransfection, the changes of MHC Ⅱexpression were detected by flowcytometry, and the mRNA abundance of CⅡTA and MHC Ⅱ was measured by FQ-PCR after inducing with recombinant rat interferon-gamma (IFN-γ). RESULTS: Different siRNA showed different reduction in MHC Ⅱ and CⅡTA expression compared with the control (P<0.01). Among the five groups, the siRNA-4 was the most efficient. The mRNA content of CⅡTA and MHC Ⅱ were reduced by 95.10%±1.25% and 82.70%±1.95% respectively and the expression of MHC Ⅱ was inhibited by over 80% in siRNA-4 group at 24 hours posttransfection. CONCLUSIONS: The special siRNA targeting to CⅡTA inhibits CⅡTA mRNA and further inhibits its regulation of MHC Ⅱ molecular expression. The blockade of MHC Ⅱ by siRNA may be useful for further studying allogeneic corneal limbal transplantation. 相似文献
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Xiaokai Song Lixin XuRuofeng Yan Xinmei HuangMohammed Ali A. Shah Xiangrui Li 《Veterinary parasitology》2009
The immunization procedure of DNA vaccine pcDNA–TA4–IL-2 of Eimeria tenella, including route, dose, time of immunization and age of primary immunization of chicken, was optimized. The stability and the cross-species protection of the vaccine were also analyzed. Efficacy of immunization was evaluated on the basis of oocyst decrease ratio, lesion score, body-weight gain and the anti-coccidial index (ACI). Chinese Yellow chickens were randomly distributed into corresponding groups (30/group). The challenged, unchallenged and vector control groups were designed. The results illustrated that 25 μg was the optimal dose and intramuscular injection was the most effective route to induce protective immunity. There were no significant differences of ACIs between boosting and non-boosting groups. Storage time and temperature had little effect on the immunizing efficacy of the vaccine. The vaccine could provide partial cross-protection against the challenge with E. necatrix and E. acervulina, but not with E. maxima. 相似文献
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提取鸡致病性大肠杆菌分离株O1、O2和O78的基因组DNA作模板,用TD-PCR技术从其中分别扩增出0.55 KB的I型菌毛结构基因(piliA).将扩增得到的piliA基因片段,用TA克隆的方法分别克隆进pGEM-T栽体中,转化至受体菌JM109中,用Amp/IPTG/X-gal琼脂平板蓝白菌落筛选法,得到舍阳性重组子的菌株,提取质粒用PstI单酶切及NcoI和PstI双酶切鉴定,结果证实,所构建的克隆质粒中均含有相应piliA基因.经DNA序列分析,其结构基因阅读框架大小为549 bp,但其中O1菌株Ⅰ型菌毛基因在第72位发生突变,有6个碱基插入.经DNAStar核酸分析软件分析,3个基因同源性为89.9%～92.0%. 相似文献
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High quality fruit production is the cornerstone of marketability. Optimum plant performance depends on the balanced and timely availability of mineral nutrients. In addition to element concentrations, the ratio of nutrient elements in solution plays a determinative role in growth, productivity, quality, and nutrients uptake. In this experiment, the effects of different Potassium:Calcium (K:Ca) ratios (1.6, 1.4, 1.2, 1, 0.85, and 0.6) in nutrient solution were studied on quality attributes of strawberry ‘Selva’. The highest and lowest leaf number and leaf area were observed in K:Ca 1.4 and 1, respectively. The highest fruit pH, electrical conductivity, total soluble solids/titratable acidity ratio, vitamin C content, ellagic acid, and color were resulted in K:Ca (1.4) ratio. K:Ca (1.6) ratio produced the highest content of protein. Moreover, K:Ca (0.85) ratio was the most effective treatment on fruit firmness. The increased quality attributes were observed in high K:Ca ratios, hence low K:Ca ratio resulted in increased fruit firmness. In conclusion, nutrient solution containing K:Ca ratios between 1 and 1.6 were suitable for producing strawberry ‘Selva’. Taken together, K:Ca (1.4) was an appropriate ratio for producing strawberry ‘Selva’ in soilless culture with coconut fiber: perlite medium. 相似文献
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三十烷醇是一种广谱性的植物生长促进剂,其促进生长效果稳定,无污染,无毒害。本文研究了不同浓度三十烷醇溶液对绿豆生长的影响,结果表明：低浓度三十烷醇溶液对绿豆生长起促进作用。 相似文献
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OBJECTIVE: The present study was conducted to examine the presence of protein kinase Calpha (PKCalpha) in the pulmonary arterioles of broilers during the development of pulmonary hypertension and pulmonary vascular remodelling. METHOD: One hundred and sixty day-old Avian-2000 broilers were divided equally into a control group and a cold temperature group. All the birds were reared in normal temperatures up to day 14, with the lighting schedule at 24 h per day. Thereafter, birds in the cold temperature group were subjected to low temperature by lowering 1-2 degrees C per day to 12-14 degrees C, and then kept constant until day 49, while birds in the control group were still brooded at normal temperatures. All the birds were fed a diet of pellets throughout the study. Samples of blood were taken from the wing vein, and of heart and lung collected after the birds were killed with an overdose of sodium pentobarbitial, at days 24, 32, 39 and 45 of age, respectively. Right ventricle to total ventricle ratio (RV/TV) and packed cell volume (PCV) were measured. Vessel wall area to vessel total area ratio (WA/TA) and mean media thickness in pulmonary arterioles (mMTPA) was examined using computer-image analytic software. Expression of PKC in pulmonary muscular arterioles was assessed by immunohistochemistry and quantified by measuring optical density (OD) using computer-image analytic software. RESULTS: The incidence of pulmonary hypertension syndrome (PHS) was 12.5% in birds exposed to cold, and 3.75% in the control group (P<0.05). PCV in the cold temperature group was elevated after day 32 (P<0.05), and RV/TV ratio increased on day 45 (P<0.05). Both the WA/TA and mMTPA of birds subjected to cold were significantly elevated (P<0.05). The OD values were not significantly increased before day 32 (P>0.05), however, one week later (at day 39 of age), the difference between the two groups was significant (P<0.05). The increased PKCalpha expression was positively correlated with the values of mMTPA and WA/TA. CONCLUSION: PKCalpha expression was up-regulated during the development of pulmonary hypertension. The activation of PKCalpha might be involved in the development of pulmonary vascular remodelling. 相似文献
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本研究以大棚延后栽培的‘南丰蜜橘’作为研究材料,结合覆盖材料与保水剂做树盘试验处理,分析不同树盘处理对浮皮程度、果实品质和树盘土壤温湿度的影响。结果表明:在果实成熟的11月,树盘覆盖处理可提升果实可溶性固形物（TSS）、可滴定酸（TA）和维生素C含量,反光膜+保水剂处理提升果实TSS和TA效果显著,分别较对照（CK）高2.30%、0.27%。果实浮皮程度与TSS、TA和维生素C含量负相关;在12月,CK和稻草处理果实TSS、TA含量低,浮皮趋于严重,浮皮程度分别高达1.67级、1.07级,而其他处理果实出现轻度浮皮,均为0.73级。浮皮程度与单果重、果实TA和维生素C含量有很大关联性;在翌年1月,浮皮程度与土壤湿度呈极显著正相关（相关系数为0.879）,而TA与土壤湿度呈极显著负相关（相关系数为-0.911）。反光膜+保水剂处理土壤湿度一直处于最高,此时期TA迅速降低,浮皮程度呈加速趋势。因此,土壤湿度越高,浮皮程度越严重,果实的TSS、TA和维生素C含量越低。通过覆盖处理能降低土壤湿度,提升果实TSS、TA和维生素C含量等品质,推迟‘南丰蜜橘’果实浮皮的发生,减轻果实浮皮程度。其中,地布覆盖处理防治果实浮皮效果最佳,反光膜、地毯、地布+保水剂处理的效果次之。 相似文献