药用植物的多倍体育种

阐述了药用植物多倍体的特征,系统地介绍了药用植物多倍体育种的方法,包括多倍体育种材料的选择方法,常用的多倍体诱导方法,多倍体材料的鉴定方法,指出了药用植物多倍体育种存在的一些问题,分析了药用植物多倍体材料的利用价值。     

Treatment and surveillance of polypoid lesions of the gallbladder in the United Kingdom

Objectives

The increase in the routine use of abdominal imaging has led to a parallel surge in the identification of polypoid lesions in the gallbladder. True gallbladder polyps (GBP) have malignant potential and surgery can prevent or treat early gallbladder cancer. In an era of constraint on health care resources, it is important to ensure that surgery is offered only to patients who have appropriate indications. The aim of this study was to assess treatment and surveillance policies for GBP among hepatobiliary and upper gastrointestinal tract surgeons in the UK in the light of published evidence.

Methods

A questionnaire on the management of GBP was devised and sent to consultant surgeon members of the Association of Upper Gastrointestinal Surgeons (AUGIS) of Great Britain and Ireland with the approval of the AUGIS Committee. It included eight questions on indications for laparoscopic cholecystectomy and surveillance based on GBP (size, number, growth rate) and patient (age, comorbidities, ethnicity) characteristics.

Results

A total of 79 completed questionnaires were returned. The vast majority of surgeons (>75%) stated that they would perform surgery when a single GBP reached 10 mm in size. However, there was a lack of uniformity in the management of multiple polyps and polyp growth rate, with different surveillance protocols for patients treated conservatively.

Conclusions

Gallbladder polyps are a relatively common finding on abdominal ultrasound scans. The survey showed considerable heterogeneity among surgeons regarding treatment and surveillance protocols. Although no randomized controlled trials exist, national guidelines would facilitate standardization, the formulation of an appropriate algorithm and appropriate use of resources.     

Extreme polyploidy in a large bacterium

Cells rely on diffusion to move metabolites and biomolecules. Diffusion is highly efficient but only over short distances. Although eukaryotic cells have broken free of diffusion-dictated constraints on cell size, most bacteria and archaea are forced to remain small. Exceptions to this rule are found among the bacterial symbionts of surgeonfish; Epulopiscium spp. are cigar-shaped cells that reach lengths in excess of 600 mum. A large Epulopiscium contains thousands of times more DNA than a bacterium such as Escherichia coli, but the composition of this DNA is not well understood. Here, we present evidence that Epulopiscium contains tens of thousands of copies of its genome. Using quantitative, single-cell PCR assays targeting single-copy genes, we have determined that copy number is positively correlated with Epulopiscium cell size. Although other bacteria are known to possess multiple genomes, polyploidy of the magnitude observed in Epulopiscium is unprecedented. The arrangement of genomes around the cell periphery may permit regional responses to local stimuli, thus allowing Epulopiscium to maintain its unusually large size. Surveys of the sequences of single-copy genes (dnaA, recA, and ftsZ) revealed genetic homogeneity within a cell consistent with only a small amount ( approximately 1%) of the parental DNA being transferred to the next generation. The results also suggest that the abundance of genome copies in Epulopiscium may allow for an unstable genetic feature, a long mononucleotide tract, in an essential gene. With the evolution of extreme polyploidy and large cell size, Epulopiscium has acquired some of the advantages of eukaryotic cells.     

Genotoxic effects of aluminum chloride in cultured human lymphocytes treated in different phases of cell cycle.

Aluminum (Al) is the most abundant metal and the third common chemical element on earth. It is known that Al is toxic, especially its trivalent form (Al(3+)), that represents the its most soluble form. Al intoxication is related to some pathogenic disorders, principally neurodegeneratives ones as Parkinson and Alzheimer diseases. The present study aimed to evaluate the mutagenic potential of aluminum chloride (AlCl(3)). Comet assay and chromosome aberrations analysis were applied to evaluate the DNA-damaging and clastogenic effects of AlCl(3), respectively, in different phases of the cell cycle. Cultured human lymphocytes were treated with 5, 10, 15 and 25 microM aluminum chloride during the G1, G1/S, S (pulses of 1 and 6h), and G2 phases of the cell cycle. All tested concentrations were cytotoxic and reduced significantly the mitotic index in all phases of cell cycle. They also induced DNA damage and were clastogenic in all phases of cell cycle, specially in S phase. AlCl(3) also induced endoreduplication and polyploidy in treatments performed during G1 phase. The presence of genotoxicity and polyploidy on interphase and mitosis, respectively, suggests that aluminum chloride is clastogenic and indirectly affects the construction of mitotic fuse in all tested concentrations.     

半夏多倍体复合体基因组DNA甲基化状态的MSAP分析

目的分析半夏多倍体复合体DNA甲基化状态的变化(甲基化水平和甲基化差异模式)。方法应用甲基化敏感扩增多态性(methylation sensitive amplified polymorphism,MSAP)技术,采用34对引物进行选择性扩增。结果扩增共得到7708条带,其中5636条带为甲基化多态性带在半夏7倍、8倍、9倍、10倍体值株间DNA甲基化水平变化不大。总扩增位点甲基化水平在54%～58%,全甲基化率为24.1%～24.3%,高倍性植株甲基化程序相应比较高。除甲基化水平稍有变化外,半夏不同倍性株间的DNA甲基化模式也形式各异,检测到的带型分为两大类(和型)。其中,不同倍性间DNA甲基化状态保持不变的位点占52.5%,归为型;少部分检测位点(占47.5%,归为型)的DNA甲基化模式在不同倍性间存在显著差异。结论半夏多倍体复合体各倍性植株间存在大量的胞嘧啶甲基化变异且整体甲基化水平较高。     

半夏多倍体复合体基因组DNA甲基化状态的MSAP分析

目的分析半夏多倍体复合体DNA甲基化状态的变化(甲基化水平和甲基化差异模式)。方法应用甲基化敏感扩增多态性(methylation sensitive amplified polymorphism,MSAP)技术,采用34对引物进行选择性扩增。结果扩增共得到7708条带,其中5636条带为甲基化多态性带在半夏7倍、8倍、9倍、10倍体值株间DNA甲基化水平变化不大。总扩增位点甲基化水平在54%～58%,全甲基化率为24.1%～24.3%,高倍性植株甲基化程序相应比较高。除甲基化水平稍有变化外,半夏不同倍性株间的DNA甲基化模式也形式各异,检测到的带型分为两大类(和型)。其中,不同倍性间DNA甲基化状态保持不变的位点占52.5%,归为型;少部分检测位点(占47.5%,归为型)的DNA甲基化模式在不同倍性间存在显著差异。结论半夏多倍体复合体各倍性植株间存在大量的胞嘧啶甲基化变异且整体甲基化水平较高。     

卵裂球电融合方法制作小鼠多倍体胚胎

目的：探索制作小鼠多倍体胚胎的方法，方法：应用卵裂球电融合方法。制作小鼠四倍体和八倍体胚胎。结果：经卵裂球电融合得到的多倍体胚胎，在体外培养条件下，可以发育到囊胚，其囊胚的形态与正常二倍体囊胚无区别，但囊胚中的细胞数量随着染色体倍性的增加而减少。进行电融合时，电压强度直接影响到卵裂球的融合效率，最适宜的电压强度与胚胎的染色体倍性存在一定的联系。结论：卵裂球电融合方法可以用来制作小鼠多倍体胚胎。     

Intrinsic karyotype stability and gene copy number variations may have laid the foundation for tetraploid wheat formation

Polyploidy or whole-genome duplication is recurrent in plant evolution, yet only a small fraction of whole-genome duplications has led to successful speciation. A major challenge in the establishment of nascent polyploids is sustained karyotype instability, which compromises fitness. The three putative diploid progenitors of bread wheat, with AA, SS (S ∼ B), and DD genomes occurred sympatrically, and their cross-fertilization in different combinations may have resulted in fertile allotetraploids with various genomic constitutions. However, only SSAA or closely related genome combinations have led to the speciation of tetraploid wheats like Triticum turgidum and Triticum timopheevii. We analyzed early generations of four newly synthesized allotetraploid wheats with genome compositions S^shS^shA^mA^m, S^lS^lAA, S^bS^bDD, and AADD by combined fluorescence and genomic in situ hybridization-based karyotyping. Results of karyotype analyses showed that although S^shS^shA^mA^m and S^lS^lAA are characterized by immediate and persistent karyotype stability, massive aneuploidy and extensive chromosome restructuring are associated with S^bS^bDD and AADD in which parental subgenomes showed markedly different propensities for chromosome gain/loss and rearrangements. Although compensating aneuploidy and reciprocal translocation between homeologs prevailed, reproductive fitness was substantially compromised due to chromosome instability. Strikingly, localized genomic changes in repetitive DNA and copy-number variations in gene homologs occurred in both chromosome stable lines, S^shS^shA^mA^m and S^lS^lAA. Our data demonstrated that immediate and persistent karyotype stability is intrinsic to newly formed allotetraploid wheat with genome combinations analogous to natural tetraploid wheats. This property, coupled with rapid gene copy-number variations, may have laid the foundation of tetraploid wheat establishment.Polyploidy or whole-genome duplication (WGD) is a driving force in plant and vertebrate evolution (1–5). However, recent molecular phylogenetic studies have argued against a general creative role of WGD in plant evolution (6–8). This incongruence in opinions is long standing; in fact Stebbins in his seminal work in 1970s stated, “polyploidy has contributed little to progressive evolution” (9). Clearly, the two schools of thought regarding the roles of WGD in plant evolution have existed for years, and can converge if we accept the idea that WGDs have occurred frequently in nature but only a small fraction thereof have contributed to successful speciation (7, 10). However, the genetic and genomic factors determining one of the two fundamental fates for a nascent polyploid remained elusive.The Triticum–Aegilops complex includes both diploid and polyploid species with phylogenetically well-defined organismal relationships (11). This plant group is therefore suitable to address the issue raised above. This is especially so because all polyploid species of the Triticum–Aegilops complex represent examples of evolutionarily recent successful speciation events via allopolyploidization, i.e., hybridization and doubling of genomes from Triticum/Aegilops species (11). Speciation of allohexaploid common wheat, Triticum aestivum L., the founder crop for the initial establishment of Middle-Eastern and European agriculture, is characterized by two sequential polyploidization events, i.e., allotetraploidization and allohexaploidization. The former event involved hybridization of two diploid species, Triticum urartu (genome AA) and a yet-unknown or extinct goat-grass species of the genus Aegilops section Sitopsis (genome SS ∼ BB). This event led to the speciation of allotetraploid emmer wheat, Triticum turgidum ssp. dicoccoides (12). The latter event that involved hybridization of a primitive domesticated form of T. turgidum (genome BBAA) with a goat-grass species Aegilops tauschii (genome DD) led to the speciation of common wheat (11). In parallel, allohexaploidization by hybridization of another allotetraploid wheat, Triticum timopheevii (genome GGAA) (13) and Einkorn wheat, Triticum monococcum (genome A^mA^m) has resulted in the speciation of Triticum zhukovskyi (genome GGAAA^mA^m) (14) (Fig. S1A). Thus, the natural hexaploid bread wheat has three diploid progenitors (13) (Fig. S1A) that have diverged from a common ancestor only about 2.5–4.5 Mya (15). Therefore, it is perhaps not surprising that allotetraploidization by hybridization of any two of these three diploid species can still be accomplished artificially to produce fertile tetraploid plants (16, 17) (Fig. S1B). Furthermore, the diploid progenitor species are known to exist sympatrically across several geographical areas in the Eastern Mediterranean region and the Near East (13). These features of the diploid progenitor species of polyploid wheat raise an intriguing question: Why has only the genome combination of SSAA or closely related ones led to successful speciation of the two natural allotetraploid wheat species, T. turgidum and T. timopheevii? This question is germane to the more general issue as to why only a small fraction of WGDs have led to successful speciation in the evolutionary history of angiosperms.A major challenge for the establishment of newly formed allopolyploids as new species is sustained karyotype instability, in particular, whole-chromosome aneuploidy, which, at the organismal level, is associated with compromised cellular metabolism and reduced fitness (18). In newly formed allohexaploid wheat, extensive whole-chromosome aneuploidy was reported (19, 20). However, the immediate chromosomal consequences associated with formation of allotetraploid wheat have remained uninvestigated.Here, we conducted in-depth molecular cytogenetic analyses of karyotype stability using fluorescence and genomic in situ hybridization (FISH and GISH) techniques and performed locus-specific molecular genetic analysis of gene copy-number variations (CNVs) in a set of protein-coding genes using newly synthesized allotetraploid wheats. Different genomic combinations of diploid Triticum/Aegilops species that are parsimoniously representing the diploid progenitors of tetraploid and hexaploid wheats were used in this study. We document that dramatic differences in karyotype stability of both chromosome number and structure existed between the S^shS^shA^mA^m/S^lS^lAA versus the S^bS^bDD/AADD genome combinations. Remarkably, localized loss or gain of DNA repeats and CNVs of gene homologs occurred widely in both chromosome stable lines, S^shS^shA^mA^m and S^lS^lAA, which, in their genomic constitutions, mimic natural tetraploid wheats, T. turgidum and T. timopheevii. We propose that persistent karyotype stability coupled with localized genomic changes and rapid CNVs of gene homologs, have laid the foundation for successful tetraploid wheat establishment and speciation.     

低剂量X射线照射对K562细胞周期及凋亡影响的实验研究

目的 低剂量照射(low-dose radiation,LDR)可调节细胞信号传导,通过各种基因和蛋白的表达改变其分子生物学效应,由此产生了多样复杂且不同于大剂量照射的生物效应.用LDR及LDR联合大剂量照射(high-dose radiation,HDR)人红白血病细胞系K562(CML),探讨LDR对K562细胞周期进程及凋亡的影响.方法 按照K562细胞照射剂量不同分为4组,对照组(0 Gy)、LDR组(0.08、0.50 Gy)、HDR组(6 Gy)及LDR联合HDR组(0.08/6 Gy、0.50/6 Gy,LDR/HDR组).采用6 MVX射线照射,LDR与HDR间隔8 h;照射后按不同时间点收集细胞,流式细胞仪检测细胞周期变化与凋亡并分析细胞DNA倍体变化.结果 LDR后6h各组S期比例增加,对照组与LDR各组分别为55.38±2.22 vs 71.91±7.30、73.13±4.09(Z=-2.428,P=0.022;Z=-3.987,P＜0.001);12 h G2/M期细胞比例增加,出现G2/M阻滞,24 h达峰值,对照组与LDR各组分别为17.81±1.27 vs 26.61±7.82、29.02±2.76(Z=-3.684,P＜0.001;Z=-2.928,P＜0.001);48 h各组G0/G1细胞增多,72 h达峰值,对照组与LDR各组分别为27.07±1.19 vs 52.32±2.42、44.06±1.90(Z=-2.351,P=0.020;Z=-2.172,P=0.032).LDR/HDR后6hS期细胞比例增多,且G2/M期比例增加,即G2/M期阻滞,12 h达峰值并持续至24 h,12 h HDR组与LDR/HDR各组G2/M期分别为26.98±2.15 vs 56.27±1.57、69.31±2.51(Z=-2.564,P=0.021;Z=-7.759,P＜0.001).LDR后48 h凋亡率增加,96 h达峰值,对照组与LDR各组分别为1.82±0.12 vs3.21±0.20、6.28±0.30(Z=-3.959,P=0.003;Z=-9.705,P＜0.001);LDR/HDR照射后24 h凋亡率增加,120 h达峰值,HDR组与LDR/HDR各组分别为14.21±0.61 vs 17.38±0.92、27.91±1.07(Z=-2.986,P=0.027;Z=-6.973,P＜0.001).LDR后6h各组四倍体及八倍体细胞增加,且随照射剂量增大而增多,对照组与LDR各组八倍体细胞分别为2.41±0.15 vs 4.84±0.46、7.83±0.59,差异有统计学意义,H值分别为5.956和7.200,P值分别为0.025和0.001;LDR/HDR后6h各组四倍体及八倍体细胞增加,亦随LDR剂量增大而增多,HDR组与LDR/HDR各组八倍体细胞分别为6.49±1.05 vs 10.80±1.23、13.77±0.79,差异有统计学意义,H值分别为5.600和7.200,P值分别为0.05和0.001.结论 LDR能诱导K562凋亡,并能增强随后HDR对K562凋亡作用;LDR能诱导S/M期解偶联及G2/M期阻滞等细胞周期进程改变;LDR诱导K562凋亡可能与S/M期解偶联及G2/M期阻滞等改变细胞周期进程有关.     

多倍体黄精中多糖和皂苷的提取及含量测定

目的：比较1年生人工栽培多倍体（四倍体）黄精与1年生人工栽培二倍体黄精、4年生野生二倍体黄精中多糖以及皂苷的含量。方法：多糖采用索氏抽提法提取,蒽酮硫酸比色法测定含量;总皂苷采用超声波提取法提取,香草醛高氯酸比色法测定含量。结果：以葡萄糖计,4年生野生二倍体黄精、1年生人工栽培多倍体黄精以及1年生人工栽培二倍体黄精中多糖含量分别为17．7％、18．6％和17．0％;以人参皂苷Rb,计,4年生野生二倍体黄精、1年生人工栽培多倍体黄精以及1年生人工栽培二倍体黄精中总皂苷含量分别为7．5％、7．5％和3．6％。结论：1年生人工栽培多倍体黄精主要活性成分（多糖和皂苷）的含量接近或略高于4年生野生二倍体黄精,皂苷含量明显高于1年生人工栽培二倍体黄精。