青葙SRAP体系的建立和优化

目的 探讨影响青葙SRAP(sequence-related amplified polymorphism)扩增的各种因素,建立并优化青葙SRAP反应体系,为分子水平鉴定青葙子及其伪品,探讨青葙不同种质问的亲缘关系奠定基础.方法 用CTAB法提取青葙DAN,设计Taq酶浓度(0.02、0.04、0.06 U/μL)、dNTP浓度(0.10、0.20、0.30 mmol/L)、Primer浓度(0.15、0.30、0.45μmol/L)3水平3因素试验和Mg2 浓度(0.5、1.0、1.5、2.0、2.5、3.0、3.5、4.0 mmol/L)8水平单因素试验.在25 μL体系中加入模板DNA 20 ng,对体系进行优化,用琼脂糖进行检测.结果 建立了适合青葙的SRAP体系,在25 μL体系中,DNA为20 ng,Taq酶浓度为0.04 U/μL,dNTP浓度为0.30 mmol/L,Primer浓度为0.30 μmol/L,Mg2 浓度为2.5 mmol/L,优化后的体系目标条带增多,且比较稳定,重现性好,得到了较好的扩增效果.结论 本研究建立的反应体系适合青葙SRAP的研究.为从分子水平鉴定青葙子正品与伪品以及鉴定青葙不同种质资源奠定了基础.     

家蝇相关序列扩增多态性扩增体系的建立与优化

目的:建立和优化家蝇的相关序列扩增多态性(SRAP-PCR)扩增体系.方法:以驯养家蝇为材料,通过单因子实验分别研究了模板DNA、Mg2+、dNTPs、引物浓度及Taq DNA聚合酶用量对家蝇SRAP分子标记扩增体系的影响,并对扩增体系进行了优化.结果:优化后的反应体系总体积为30 μl,含50 ng模板DNA、1.5mmol/L Mg2+、0.20 mmol/L dNTPs、0.15 μmol/L引物和0.50 U Taq DNA聚合酶.结论:家蝇SRAP-PCR扩增优化系统的建立,为今后利用SRAP技术进行家蝇遗传多样性分析、基因定位奠定了基础.     

环带库蚊ISSR-PCR反应体系的建立及优化

目的:建立及优化适宜环带库蚊(Culex annulus)ISSR-PCR的反应体系,为环带库蚊遗传多样性研究奠定技术基础.方法:选用引物IS01,应用正交试验及单因素试验对的ISSR-PCR反应体系中的BSA、模板DNA、Mg2+、引物、dNTP、Taq DNA聚合酶浓度6个关键因素进行优化筛选.结果:20 μL反应体系中最佳因素为BSA 2.0 g/L、DNA模板1.00×10-4mg/L、Mg2+2.0 mmol/L、引物1.0 μmol/L、dNTP0.15 mmol/L、TaqDNA聚合酶0.20 5U/μL.结论:优化后的反应体系适于环带库蚊的ISSR-PCR扩增.     

小鼠RAPD-PCR反应体系及扩增程序的筛选

目的:筛选优化小鼠RAPD-PCR反应体系及扩增程序.方法:以40条引物对9个品系的小鼠基因组DNA在各种不同条件下进行RAPD-PCR,分析比较扩增结果.结果:在下列反应体系及扩增程序下结果较好:在25 μl的反应体系中含有10×buffer 2.5 μl,dNTP 200μmol/L, Mg2+ 2.5 mmol/L,引物0.2 μmol/L,Taq酶1.5 U,模板DNA 20 ng,扩增程序为94℃变性1 min,38℃退火1 min,72℃延伸2 min,40个循环后接10 min延伸.结论:以上述反应体系及扩增程序进行小鼠RAPD-PCR,扩增结果稳定.     

白芨ISSR-PCR反应体系的建立及优化

目的：建立并优化白芨ISSR-PCR反应体系,为白芨种质资源分子评价奠定技术基础.方法：应用单因素和正交试验研究ISSR-PCR反应体系中模板DNA、Mg2、引物、dNTP、BSA及Taq DNA聚合酶等6个主要成分对扩增结果的影响,优化出ISSR-PCR最佳反应体系.结果：优化后25 μL反应体系中含100 ng DNA模板、2.0 mmol/L Mg2＋、2.5 μmol/L引物、0.2 mmol/L dNTP浓度、3.75 g/L BSA及1.25 UTaq DNA聚合酶.结论：建立了适用于白芨的ISSR-PCR最佳反应体系.     

中金钗石斛相关序列扩增多态性反应体系的正交优化研究药大质量观及实践

目的 建立金钗石斛Dendrobium nobile相关序列扩增多态性 (SRAP) 反应体系,为金钗石斛分子生物学研究提供技术支持。方法 运用 L25(56) 正交设计对影响金钗石斛 SRAP 反应的5个因素：模板 DNA、Mg2+、dNTPs、Taq 酶、引物在5个水平上进行优化试验,对 PCR 结果进行极差分析。结果 金钗石斛 SRAP 反应最佳体系为：25 μL PCR 体系中含有 20 ng 模板 DNA,2.0 mmol/L Mg2+,0.2 mmol/L dNTPs, 1 U Taq 酶,0.6 μmol/L 引物。各因素对 SRAP 反应的影响依次为：〖WTBX〗Taq 酶＞Mg2+＞dNTPs＞模板 DNA＞引物。结论 所建立的金钗石斛 SRAP 反应体系具有标记位点清晰、多态性丰富、稳定性好等特点,可用于金钗石斛分子生物学的研究。     

正交设计优化脑膜炎奈瑟氏菌PCR反应体系

目的:建立脑膜炎奈瑟氏菌PCK检测的最佳反应体系,探讨影响检测的多个因素.方法:利用正交设计,对膜炎奈瑟氏菌PCR反应体系的4因素(Taq DNA聚合酶、引物、Mg2 、dNTP)在3个水平上进行优化实验,筛选出各反应因素的最佳水平.结果:正交实验结果表明,最佳反应体系为(50μl):Taq DNA聚合酶2.0U,引物0.3μmol/L,Mg2 3.0mmol/L,dNTP100μmol/L.结论:所得体系反应稳定,正交法实验设计高效、快捷,科学性强,值得推广.     

夏枯草ISSR分子标记技术的建立与体系优化

目的 建立并优化夏枯草ISSR-PCR反应体系和扩增程序,为探讨夏枯草种质间遗传多样性奠定基础。方法 采用单因子试验和正交设计方法,研究Mg2+、dNTP、引物、 Taq DNA聚合酶、模板DNA、退火温度及循环次数对PCR扩增的影响。结果 夏枯草ISSR-PCR的最佳反应体系为：在20 μL的反应体系中含模板DNA 30 ng,Mg2+2.2 mmol/L、dNTP 175 μmol/L、引物0.75 μmol/L、Taq DNA聚合酶1.0 U。在此基础上,从92条引物中筛选出18条扩增稳定、多态性丰富的ISSR引物,并通过梯度PCR试验,确定引物最佳退火温度。结论 采用单因子试验和正交设计方法可以快速建立ISSR-PCR反应体系,经过24份夏枯草种质检验,证明该体系稳定可靠,可用于夏枯草遗传分析。     

黄花蒿SRAP-PCR反应体系的建立与优化

目的用序列相关扩增多态性(SRAP)这一新的分子标记技术建立稳定可重复的黄花蒿Artemisia annua的SRAP最佳反应体系。方法以黄花蒿DNA为模板,分析了SRAP反应体系中的一些重要参数对PCR扩增的影响。结果建立了稳定可重复的SRAP反应体系;25μL反应体系中,模板DNA为30～105ng,MgCl2为2.0mmol/L,dNTP为0.25mmol/L,Taq酶为1.0U,引物为2.0μmol/L;扩增程序:94℃预变性3min,94℃变性1min,35℃复性1min,72℃延伸1min,5个循环;94℃变性1min,50℃复性1min,72℃延伸1min,35个循环,72℃延伸7min。结论本研究建立的黄花蒿SRAP体系重复性好,稳定性强。     

大黄ISSR-PCR反应体系的建立与优化

采用正交设计和单因素试验相结合的方法对影响大黄ISSR-PCR反应体系的重要因素(Mg2+、Taq DNA聚合酶、dNTPs、引物、DNA模板)进行优化,通过统计软件分析和直观分析获得最佳的反应体系:DNA模板30 ng,2.5 μL 10×Taq Buffer,dNTPs 0.4 mmol/L、引物0.25 μmol/L、Mg2+ 2.5mmol/L、Taq DNA聚合酶1.25 U.     

Value of D-Dimers in patients with acute aortic dissection

Objective: To evaluatel the value of D-dimers in patients with acute aortic dissection (AAD). Methods: This study consisted of 16 patients with AAD and 27 non-AAD patients. Serum D-dimets were measured by Sta-Liatest D-DI immunoturbidimetric assay. Results: D-dimer level was higher (P ＜ 0.001) in patients with AAD(7.91 ± 5.52 μg/ml) than that in non- AAD group(1.57±1.24 μg/ml). D-dimer was positive (＞0.4 μg/ml) in all patients with AAD and in 10 control group patients (37%). Among patients with acute AAD, D-dimers tended to be higher in Stanford A than in Stanford B (8.67 ± 4.31 μg/ml vs. 3.24±1.27 μg/ml, P ＜0.01). D-dimer values tended to be higher in more extended disease(3.84 ± 1.65 μg/ml, 8.57 ± 3.58 μg/ml and 11.87 ± 5.69 μg/ml in thoracic aorta, thoracic and abdominal aorta, thoracic and abdominal aorta and iliacal arteries, respectively, P ＜ 0.05 for both 8.57 ± 3.58 and 11.87 ± 5.69 vs. 3.84 ± 1.65 ). Including the control group into the analysis, we found a sensitivity of 100%, a negative predictive value of 100%, and a specificity of 66% and a positive predictive value of 64% for D-dimer in diagnosis of AAD in our patients with suspected AAD. Conclusion: D-dimer was elevated in patients with AAD. A negative D-dimer test result could be useful in excluding AAD.     

Research of combined liver-kidney transplantation model in rats

Objective： To set up a simple and reliable rat model of combined liver-kidney transplantation. Methods： SD rats served as both donors and recipients. 4℃ sodium lactate Ringer＇s was infused from portal veins to donated livers,and from abdominal aorta to donated kidneys, respectively. Anastomosis of the portal vein and the inferior vena cava （IVC） inferior to the right kidney between the graft and the recipient was performed by a double cuff method, then the superior hepatic vena cava with suture. A patch of donated renal artery was anastomosed to the recipient abdominal aorta. The urethra and bile duct were reconstructed with a simple inside bracket. Results： Among 65 cases of combined liver-kidney transplantation, the success rate in the late 40 cases was 77.5%. The function of the grafted liver and kidney remained normal. Conclusion： This rat model of combined liver-kidney transplantation can be established in common laboratory conditions with high success rate and meet the needs of renal transplantation experiment.     

安心颗粒对急诊经皮冠状动脉介入术后生活质量影响的研究

目的：评价使用安心颗粒对急诊经皮冠状动脉介入术（PPCI）术后生活质量的影响.方法：将160例接受PPCI的急性ST段抬高型心肌梗死患者随机分为安心颗粒组（术前顿服安心颗粒8.8g,术后安心颗粒4.4 g/次,每日2次）和对照组（仅接受基础药物治疗）.所有患者均服用阿司匹林、氯吡格雷和阿托伐他汀.分别在入院时、出院前1d、出院后180 d时,应用心肌梗死多维度量表（MIDAS）、中文版SF-36评价量表对患者生活质量评分.并观察术后30 d以内的出血并发症、血小板减少症发生情况.结果：入院时和出院前1d,两组患者的心肌梗死MIDAS、SF-36量表评分比较无差异（P＞0.05）;出院后180 d时,与对照组比较,安心颗粒组MIDAS、SF-36评分明显减低（P＜0.05）;组内与入院时比较,两组出院前1d、出院后180 d时,MIDAS、SF-36评分均降低（P＜0.05）.两组患者在随访期间均无大量出血、少量出血、重度和极重度血小板减少症发生,安心颗粒组有4例、对照组有7例发生不明显出血（P＞0.05）.两组发生轻度血小板减少症的患者数比较无差异（P＞0.05）.结论：PPCI使用安心颗粒,能改善急性ST段抬高型心肌梗死患者的生活质量,且不增加出血风险.     

The comparative studies of the influences of Urapidil and Nicardipine on sino-atrial node function, atrio-ventricular node function and hemodynamics

Objective:To investigate the influences of urapidil and nicardipine on rabbit sinus function,atrio-ventricular node function and hemodynamics.Methods:Thirty-two Angora's rabbits were selected and randomly divided into four groups.U1 group:urapidil 0.25 mg/kg;U2 group:urapidil 0.5 mg/kg;N1 group:nicardipine 10 μg/kg;N2 group:nicardipine 20 μg/kg.All these medicine were administrated within 30 seconds.Measurements were taken before and after the administration of urapidil or nicardipine for the following data:mean blood pressure(MAP),heart rate(HR),sino-atrial conduction time(SACT),maximal sinoatrial recovery time(SNRTmax)corrected sinus node recovery time(CSNRT),index of sinus node recovery time(SNRTI),Wenckebach A-V conduction frequency (WB),and P-R interval.Results:Significant MAP and HR changes were identified in all of the four groups before and after administration of both urapidil and nicardipine.No significant changes could be found in the rest of the parameters.Intergroup analysis showed that SACT and CSNRT of N1 and N2 groups were shorter than those of the U2 group(P＜0.01);the MAP decreased(P＜0.01)and the HR increased drastically(P＜0.01).Conclusions:Neither urapidil(0.25 mg/kg,0.5 mg/kg)nor nicardipine(10μg/kg,20μg/kg)has any significant influence on rabbit sinus function or rabbit atrio-ventricular node function.Nicardipine could be a better choice than urapidil for parafunctional sinus node patients.     

Expression of OPGmRNA and ODFmRNA in the patients of hip fracture due to osteoporosis

Objective：To investigate the gene expression of osteoprotegerin（OPG） and osteoclast differentiation factor（ODF） in the bone tissue of patients with hip fracture due to osteoporosis. Methods：OPGmRNA and ODFmRNA in the bone tissue in 50 cases of osteoporosis sufferers（over 50 years old） with hip fracture（Observer Group） and 30 cases of hip facture sufferers with no osteoporosis（Control group） were analyzed with the Semi-Quantitative RT-PCR method. Results：The mRNA expressed of ODF, OPG were both high in the patients with hip fracture. In the control group, the expression of OPG mRNA was observed, while the expression of ODF mRNA was very slight. Conclusion：Aged patients contained all signals including OPG, ODF that are essential for inducing osteoclastogenesis and promoting bone resorption.     

The clinicopathological and immuohistochemical analysis of solid-pseudopapillary tumor of the pancreas： report of 9 cases

Objective：To investigate the clinical features, pathological characteristics and immunophenotype of solid-pseudopapillary tumor of the pancreas（SPTP）. Methods：Nine surgically treated cases of SPTP were retrospectively reviewed. Hematoxylin and Eosin（HE） staining and immunohistochemical staining were used to analyze all cases, and the general clinical data was collected. Results：Six patients were asymptomatic except for a palpable mass. Two patients complained of vague-epigastric pain. One patient appeared jaundice. The tumor was encapsulated and solid tissues alternately with cystic tissues. Histologically, the histological structure of solid portion was pseudopapillary with a fibrovascular core. Tumor cells were uniform and medium-sized which were arranged in sheets ets or nests or pseudopapillary patterns. Immunohistochemical studies demonstrated that SPTP proved positive in vimentin（9/9 cases）, AAT（9/9 cases）, NSE（9/9 cases）, ACT（7/9 cases）, CK20（2/9 cases）, CgA（1/9 cases）, S-100（3/gcases）, PR（4/gcases）, Syn（3/9 cases） and CD56（5/9cases）, negative in CEA and ER. Conclusion：SPTP is a tumor predominantly occurring in young women frequently without special symptoms. This tumor has various characteristical histological patterns with different immunophenotype.     

Electro-Acupuncture Combined with the Trigger Point Needle-Embedding for Treatment of Primary Trigeminal Neuralgia in 31 Cases

In recent years, the author of this essay has applied electro-acupuncture combined with the trigger point needle-embedding for treatment of primary trigeminal neuralgia in 31 cases, yielding satis- factory results as reported in the following.     

Investigation of Matrix Metalloproteinase -1, 2 and tissue inhibitor of matrix metalloproteinases-1 in Endometriosis

Objective： To explore the role of matrix metalloproteinase-1,2 （MMP-1, MMP-2） and tissue inhibitor of matrix metalloproteinases-1 （TIMP-1） in endometriosis. Methods： The eutopic and ectopic endometria from 40 subjects suffering from endometriosis and regular.endometria from 40 subjects （excluding endometriosis） were collected and examined by in situ hybridization technology and western blot assay. Results： Both expressions of MMP-1 and -2 were stronger in ectopic endometrium and eutopic endometrium than in normal endometrium. On the contrary, the expression of TIMP-1 in ectopic endometrium and eutopic endometrium was lower. The differences were significant （P 〈 0.01 ）. Moreover, there was no relationship among the expressions of MMP-1, 2 and TIMP-1 in ectopic endometrium. Conclusion： The expressions of MMP-1, 2 and TIMP-1 lose balance and lack of periodic changes in ectopic endometrium , which explains the biological invasive behavior of endometriosis. It was suggested-that regulating the balance between the MMPs and TIMP-1 should be an ideal therapeutic target to endometriosis.     

Shi Da-zhuo (史大卓) —An outstanding cardiologist of integrative medicine

Prof. SHI Da-zhuo, Ph.D., male, was born on March 20, 1960. Prof. SHI entered the Ph.D. program in 1990 at the China Academy of Chinese Medical Sciences under the supervision of Prof. CHEN Ke-ji, majoring in the treatment of cardiovascular diseases. After receiving his Ph.D. degree in 1993, Prof. SHI started working at the Cardiovascular Center in Xiyuan Hospital affiliated to China Academy of Chinese Medical sciences.     

Persist in and Carry Forward the Integrative Medical Research——Speech at the 6th National General Congress of Chinese Association of Integrative Medicine by Academician CHEN Zhu,Minister of Ministry of Health

The 6th National General Congress of Chinese Association of Integrative Medicine （CALM） was convened at 19-20, April 2008 in Beijing. Academician CHEN Zhu, the minister of Ministry of Health indicated at the congress that the integration of Chinese and Western medicine is very well in keeping with the situation of our country and the general rule of development in medical science; and as a good integration of Chinese medicine and Western medicine, it is mutually beneficial and advantageous to both of them. Seeing the creativity shown in integrative medical investigation in theoretic and methodological sides, we should and must persist in and develop it.