散发性结直肠癌遗传标识物研究进展

散发性结直肠癌（sporadic colorectal cancer,sCRC）是一类没有明显家族遗传倾向,由遗传因素和环境因素共同作用的消化道恶性肿瘤。近年来,分子技术的发展使发现复杂疾病潜在的生物标识物成为可能。基于全基因组关联研究（genome-wideassociationstudy,GWAS）以及高通量测序技术的应用,已经从全基因组范围内确定了大量肿瘤易感基因以及其后天突变标识物。本文对近年来散发性结直肠癌（sCRC）遗传标识物最新研究进展作一综述,并对其未来研究方向加以展望。     

结直肠癌表观遗传学研究进展

近年来,结直肠癌（colorectal cancer,CRC）成为影响人类健康的常见恶性肿瘤之一。目前对CRC发病机制的研究不仅停留在对核基因组的研究,更多集中在表观遗传学对疾病的影响,如DNA的甲基化、组蛋白乙酰化、基因印迹等对疾病发生发展过程的影响。本文主要讲述了CRC的表观遗传学研究进展,旨在探讨表观遗传学在CRC风险预警和治疗中的潜在作用。     

樱桃谷鸭源鹅出血性多瘤病毒(GHPV)全基因组克隆与分析

鹅出血性多瘤病毒(Goose hemorrhagic polyomavirus,GHPV)为近年来我国鸭群新发病原。为明确我国鸭源GHPV基因组特征,本研究利用分段PCR扩增技术克隆获得樱桃谷鸭(Anas platyrhynchos domestica)源GHPV(FJ201601株)全基因组序列,并对其进行核苷酸同源性比较和遗传进化分析。结果表明,克隆的GHPV(FJ201601株)全基因组序列长为5 254 bp(Gen Bank登录号MG544856),(G+C)%含量为48.01%,编码的6个主要功能基因(ORF-X,VP2,VP3,VP1,Large T和Small T)长度分别为510、981、654、1062、1 911和483 bp。核苷酸同源性分析表明,FJ201601株和其他不同地区、不同宿主来源GHPV相互之间基因组核苷酸同源性较高(99.7%以上);其6个主要功能基因和其他不同地区、不同宿主来源GHPV相应基因的核苷酸(99.4%以上)和氨基酸同源性(98.2%以上)均极高。从全基因组及其编码的6个主要功能基因遗传进化关系可见,GHPV各分离株在基因组上亲缘关系较近,6个主要功能基因在遗传进化上差异不大,但北京鸭源GHPV和樱桃谷鸭源GHPV在遗传进化上仍存在细微差别,且不同宿主(多品种鸭和鹅)源GHPV在遗传进化上不存在宿主特异性。本研究丰富了GHPV分子流行病学数据,明确了GHPV基因组分子特征,为后续开展GHPV相关研究提供参考。     

可供利用的植物基因克隆技术

植物基因工程研究的首要目标是把特定的靶基因从植物基因组或其它生物基因组中分离出来，只要把基因分离并克隆，才有可能了解其遗传信息，然后进行分子操作。本文主树常用图谱克隆基因技术及克隆基因；转座子标鉴克隆基因技术及克隆基因；文库克隆基因技术；基于PCR技术克隆基因；人工合成基因技术进行了综述。     

利用BSA-Seq方法快速定位作物农艺性状QTL/基因概述

为了利用生物学技术和手段来解决农业问题,让基因组测序技术、转基因及基因编辑等现代分子生物学技术为农业遗传育种服务,实现常规育种与分子设计育种的紧密结合,加速作物精准育种进程,近年颇受学术界关注。现梳理了一些分子生物学专用名词概念,概述了目前在正向遗传学研究中如何利用极端表型材料基于全基因组测序的BSA-Seq方法(MutMap法、MutMap+法、MutMap-Gap法、QTL-seq法)快速定位重要农艺性状的QTL/基因,为快速定位和克隆候选基因提供新思路。     

荷斯坦牛DRA基因多态性及序列特征分析

为探讨荷斯坦牛DRA基因的多态性及序列特征,本研究采用基因组DNA混合池扩增并直接测序的方法对荷斯坦牛DRA基因编码区进行多态性分析,同时利用生物信息学方法预测DRA基因的理化特性及其编码蛋白的高级结构。结果表明,荷斯坦牛DRA基因编码区上有4个碱基突变,包含1个错义突变和3个同义突变。DRA基因共编码253个氨基酸,编码产物是一种稳定的不可溶蛋白,具有15个潜在的磷酸化位点,二级结构为混合型,三级结构由α-螺旋、β-折叠、β-转角和无规则卷曲组成。本研究结果为荷斯坦牛的疾病相关基因筛选和抗病分子育种提供了一定的理论依据。     

经高赖氨酸基因导入玉米自交系的研究

通过对18个玉米自交系的组织培养,筛选到8个可以正常分化的自交系.用基因枪的方法对其中的4个自交系进行了遗传转化,将高赖氨酸基因导入玉米的胚性愈伤组织.以减少潮霉素筛选次数和筛选浓度的方法,得到分化再生植株.经分子检测,证明外源基因已成功整合到玉米基因组中.部分T0代植株正常结实,通过对T1代植株的分子检测,表明外源基因可以稳定遗传.对其中的转化植株所结的种子进行了赖氨酸含量的检测,发现赖氨酸含量最高提高了16%.     

百脉根BIO和豌豆突变位点ELE2的比较基因组定位

豆科两侧对称花的花瓣具有背腹（DV）的分化以及可变的器官内部（IN）非对称性,在大小与形状上显示出不同的发育特征;因而花瓣的发育为克隆决定植物器官的形状与大小的关键基因提供了很好的实验系统。本研究对百脉根中BIO基因进行研究。百脉根bio突变体具有多效性,既影响花器官内部的对称性也影响器官的大小和育性,豌豆ele突变体的表型与bio相似。定位结果表明BIO和ELE2位于豆科基因组的共线性区段,提示BIO和ELE2可能是同源基因突变所致。本研究利用比较基因组定位方法,将BIO和ELE2候选基因锚定在豆科模式植物百脉根和蒺藜苜蓿基因组含有11个同源基因的BAC重叠群上。BIO和ELE2基因的克隆将有助于揭示豆科花瓣形态和大小调控的分子机理,进而为豆科作物遗传改良提供分子理论基础。     

北极狐PMEL基因启动子活性及转录调控区域研究

前黑素小体蛋白基因(premelanosome protein gene,PMEL)可以直接启动黑素小体内纤维的形成,促进黑素小体合成,是调控动物毛色的主要候选基因之一。目前关于北极狐(Vulpes lagopus)毛色差异表达基因的相关研究相对较少,本研究旨在筛选调控北极狐毛色的PMEL基因的启动子核心区域及转录因子。研究通过基因组测序技术,获得了北极狐PMEL基因启动子序列,利用生物信息学方法对其核心启动子区域和转录因子结合位点进行预测;以北极狐基因组DNA为模板,扩增出该基因不同长度的启动子缺失片段,并连接至p GL3-Basic载体,将重组质粒瞬时转染到A375和293T细胞,利用基因检测仪进行活性验证。结果表明,成功构建了8个含有不同长度启动子片段的重组质粒,经双荧光素酶活性检测发现北极狐PMEL基因的-1 162/+8区域为核心启动子区域;生物信息学预测分析发现在北极狐PMEL基因的-1 162/-655区域存在5个转录因子结合位点,分别为Sp1(-966/-952)、Sp1(-912/-900)、Sp1(-750/-736)、NF-1(-712/-699)和NF-1(-682/-673);利用重叠延伸PCR技术成功构建了5个突变载体,经双荧光素酶活性检测发现该5个突变载体活性均显著下降(P0.05),表明这5个转录因子是北极狐PMEL基因转录调控的正调控元件。本研究为探究该基因的表达调控机制提供了科学依据,并为北极狐毛皮品质分子育种和彩色毛皮材料的创制提供了思路。     

逆转座子对真核生物基因组及宿主基因表达的影响

逆转座子广泛存在于真核生物基因组中,由于其自身结构及转座的特殊性,它们对真核生物的基因及基因组的大小、结构、功能和进化都有重要影响,尤其是在基因表达调控上,逆转座子插入能够改变宿主基因(即被插入基因)转录的时间和空间模式,造成选择性拼接、形成多种转录本等多种效应。同时,逆转座子活动所造成的稳定变异为选择新的、有突破性的优良性状提供了丰富的素材,显示出其在动植物品种改良中的巨大应用潜力,用逆转座子作基因标签进行基因功能研究、开发一些新型分子标记等都已得到广泛的应用。因此,关于真核生物中的逆转座子及其对宿主基因表达影响的研究具有十分重要的理论意义和巨大的应用前景,是目前国际上生命科学领域研究的热点课题之一,本文就该领域近年来的研究进展作一概述。     

In vitro antiproliferative activity of isothiocyanates and nitriles generated by myrosinase-mediated hydrolysis of glucosinolates from seeds of cruciferous vegetables

A comparison of the effect of isothiocyanates and nitriles derived from some glucosinolates, namely, epi-progoitrin, sinalbin, glucotropaeolin, glucocheirolin, and glucoraphenin, on human erythroleukemic in vitro cultured cells was studied. Many studies have in fact evidenced that a consumption of vegetable containing glucosinolates could reduce the development of colorectal cancer. In the experimental conditions used, the production of isothiocyanates and nitriles from glucosinolates is almost quantitative as confirmed by HPLC or GC-MS analysis. The obtained results demonstrated that in general nitriles are considerably less potent than the corresponding isothiocyanates in inhibiting cancer cell growth. Particularly, the isothiocyanates inhibitory activity on K562 cells growth is higher in the case of products derived from epi-progoitrin, glucotropaeolin, glucoraphenin, and glucocheirolin; while for nitriles the higher activity in inhibiting K562 cells growth is showed by sinalbin-derived product. Considering the antiproliferative activity found for isothiocyanates and nitriles, further studies will be aimed to the possible application of glucosinolate-derived products as chemopreventive cancer agents for the reduction of colorectal cancer.     

茂原链霉菌DSM 40847全基因组测序及序列分析

茂原链霉菌（Streptomyces mobaraensis）是一种高GC含量的革兰氏阳性放线菌,它广泛用于生产转谷氨酰胺酶、博来霉素、莫能霉素等。目前,还没有相关研究报道茂原链霉菌的全基因组序列,这限制了基于功能基因挖掘和利用的定向育种以及分子遗传研究。本研究首次通过高通量测序技术对茂原链霉菌DSM40847进行全基因组测序,使用Velvet软件进行拼接得到266 contigs,整个基因组大小约7.5 Mb,GC含量为73.2%,序列已提交至美国国立生物技术信息中心（NCBI）的GenBank数据库。通过对基因组序列进行分析,成功鉴定转谷氨酰胺酶酶原激活相关蛋白基因,同时对该菌株的次级代谢产物合成基因簇进行了预测,相关研究结果将为茂原链霉菌的功能基因组学研究提供基础数据。     

禽呼肠孤病毒分子生物学研究进展

本文就十几年来有关呼肠孤病毒的特性、基因组、基因重组、病毒蛋白、诊断方法等分子生物学的研究进行了综述,并指出了禽呼肠孤病毒分子生物学研究中存在的问题,及进一步研究的方向。     

Fruit and vegetable consumption and risk of colorectal cancer in Japan: The Miyagi Cohort Study

OBJECTIVE: Adequate fruit and vegetable intake has been suggested to protect against colorectal cancer. However, several recent prospective studies have reported no association. We therefore examined the association between fruit and vegetable intakes and the risk of colorectal cancer in a prospective cohort study in Japan. DESIGN: Between June and August 1990, 47 605 Japanese men and women completed a self-administered questionnaire, including a food-frequency questionnaire. We divided the subjects into quartiles based on their self-reported fruit and vegetable consumption. There were 165 colon cancer and 110 rectal cancer incidences identified during 7 years of follow-up, to the end of December 1997. We used Cox proportional hazards models to estimate the relative risk (RR) of developing colorectal cancer according to the level of fruit and vegetable consumption, applying adjustments for potential confounders. RESULTS: No statistically significant association was observed between fruit and vegetable consumption and the risk of colorectal cancer. The multivariate RR of colon cancer in the highest quartile of fruit and vegetable intake compared with the lowest was 1.13 (95% confidence interval (CI) 0.73-1.75), the RR for vegetables alone was 1.24 (95% CI 0.79-1.95) and that for fruit alone was 1.45 (95% CI 0.85-2.47). The corresponding multivariate RRs for rectal cancer were 1.12 (95% CI 0.67-1.89), 1.14 (95% CI 0.67-1.93) and 1.41 (95% CI 0.73-2.73). CONCLUSIONS: We found no association between the consumption of fruit and vegetables and the risk of colorectal cancer among the Japanese population.     

High molecular weight coffee melanoidins are inhibitors for matrix metalloproteases

High molecular (above 10 kDa) melanoidins isolated from coffee beans of varying roasting degree were found to be efficient inhibitors for the zinc-containing matrix metalloproteases MMP-1, MMP-2, and MMP-9 with IC(50) values ranging between 0.2 and 1.1 mg/mL in vitro. The inhibitory potential increased with roasting degree. No or only slight inhibition of other zinc-containing peptidases closely related to MMPs, namely, Clostridium histolyticum collagenase and angiotensin converting enzyme, was found, indicating specific structural features of melanoidins to be responsible for the interaction with MMPs. A continuous increase on the apparent molecular weight of melanoidins as well as incorporation of phenolic substances into the melanoidin structure with progress of roasting was observed, concomitant with a significant increase in the carbon/nitrogen of the melanoidins. This suggests that the melanoidins are mainly formed by incorporation of carbohydrates and phenolic compounds onto a proteinaceous backbone. As MMP-1, MMP-2, and MMP-9 play a pivotal role in pathogenesis of colorectal cancer, studies on possible physiological effects of melanoidins are mandatory.     

Dietary fiber intake in two European diets with high (copenhagen, Denmark) and low (Murcia, Spain) colorectal cancer incidence

Contradictory results have been reported in studies associating dietary fiber (DF) intake and the incidence of colorectal cancer. Most studies focused mainly on the amount of total DF, but DF is a heterogeneous complex and its components are generally ignored. The aim of this work was to compare the amount and composition of DF intake in Murcia (Spain) and Copenhagen (Denmark). Total dietary fiber intake was a 31% higher in Murcia than in Copenhagen, the ratio of insoluble DF/soluble DF being similar in the two diets. DF in Murcia contains major amounts of nonstarch polysaccharides and polyphenolic compounds, mainly derived from the high consumption of fruits and vegetables (6.7 servings). Resistant starch intake was of the same order in the two regions while resistant protein intake was higher in Murcia. The quantitative and qualitative differences in DF intake may contribute to understand the differences in colorectal cancer incidence in Mediterranean and Scandinavian areas.     

Cancer chemopreventive effects of lycopene: suppression of MMP-7 expression and cell invasion in human colon cancer cells

Clinical studies indicate that high blood levels of leptin or matrix metalloproteinase-7 (MMP-7; matrilysin) proteins are associated with tumor progression of human colorectal cancer (CRC). Leptin could play an important role in cell migration and invasion of cancer cells. Our previous study indicated that lycopene could inhibit the proliferation of human colon cancer cells in vitro. However, the inhibitory effects of lycopene on the progression of human colon cancer cells have not been demonstrated yet. In this study, we investigated the inhibitory effects of lycopene on tumor progression including cell invasion and MMP-7 expression in leptin-stimulated human colon cancer cells in vitro. Our results demonstrated that lycopene significantly inhibited leptin-mediated cell invasion and MMP-7 expression in human colon cancer HT-29 cells. Lycopene could augment the expression and stability of E-cadherin proteins. Our results showed that MAPK/ERK and PI3K/Akt signaling pathways played important roles in leptin-mediated MMP-7 expression and cell invasion. Lycopene could effectively inhibit the phosphorylation of Akt, glycogen synthase kinase-3β (GSK-3β) and ERK 1/2 proteins. The molecular mechanisms of lycopene were in part through decreases in nuclear levels of AP-1 and β-catenin proteins. These novel findings suggested that lycopene could act as a chemopreventive agent to suppress MMP-7 expression and leptin-mediated cell invasion in human colon cancer HT-29 cells.     

Curcuminoids-cellular uptake by human primary colon cancer cells as quantitated by a sensitive HPLC assay and its relation with the inhibition of proliferation and apoptosis

Curcumin, which is a bright orange-yellow pigment of turmeric with antioxidant properties, has been shown to produce a potent preventative action against several types of cancers in recent studies. It has also been reported to protect the development of colon tumor in animals being fed with carcinogen. In the colon cancer cells, curcumin was illustrated to inhibit cell proliferation and induce apoptosis. As an antioxidant, it acts as an anti-inflammatory as well as an antitumor agent. Curcumin has been detected to exist in nature in the form of curcuminoids, a mixture of curcumin, the major component, with two of its related demethoxy compounds (demethoxycurcumin and bisdemethoxycurcumin). In the present study, we have investigated the antiproliferation and induced apoptosis effects of curcuminoids on colon cancer, using the primary cancer cells isolated from Taiwanese colon cancer patients as the model for colorectal cancer. Results showed that curcuminoids inhibited cell proliferation and induced apoptosis of these human primary colon cancer cells. The effects were observed in a dose-dependent manner as dose increased from 12.5 to 100 microM. With the aim of furthering the fundamental understanding of the mechanisms underlying the antiproliferation and induced apoptosis effects of curcuminoids on these human colon cancer cells, we developed a sensitive, rapid, and reproducible assay method based on high-performance liquid chromatography (HPLC). This HPLC technique developed was found to successfully determine, in a quantitative manner, the cellular uptake of curcuminoids. The uptake of these curcuminoids by the colon cancer cells was shown to increase as the dose of curcuminoids was increased. The observations of inhibited proliferation and increased apoptosis in the colon cancer cells appeared to be associated with the cellular uptake of curcuminoids.     

Green and black teas inhibit atherosclerosis by lipid, antioxidant, and fibrinolytic mechanisms

Tea is the most widely consumed beverage in the world, second only to water. Most laypersons and scientists believe that green tea is healthier than black tea due to the low incidence of heart disease and cancer in the Orient. Here, we report the first dose-response comparison of a green and black tea on normal hamsters after long-term supplementation and on a hamster model of atherosclerosis. Both teas were equally effective in inhibiting atherosclerosis with the lower dose decreasing it 26-46% and the high dose decreasing it 48-63%. Atherosclerosis was inhibited by three mechanisms: hypolipemic, antioxidant, and antifibrinolytic. There was a significant correlation between atherosclerosis and the three mechanisms. In the normal animals, teas also caused some improvement in plasma low density lipoprotein (LDL), LDL/high density lipoprotein ratio, triglycerides, lipid peroxides, lower density lipoprotein lipid peroxides, and fibrinogen. Isolated lower density lipoprotein oxidizability was also reduced in all groups. Green and black teas were equally effective at human equivalent doses, thus confirming human intervention and epidemiology studies and providing mechanisms for teas' benefit.