A comparison of the genetic diversity in Dipteronia sinensis Oliv. and Dipteronia dyeriana Henry

Dipteronia is an endemic genus to China and includes only two species, Dipteronia sinensis and D. dyeriana. Based on random amplified polymorphic DNA (RAPD) markers, a comparative study of the genetic diversity and genetic structure of Dipteronia was performed. In total, 128 and 103 loci were detected in 17 D. sinensis populations and 4 D. dyeriana populations, respectively, using 18 random primers. These results showed that the proportions of polymorphic loci for the two species were 92.97% and 81.55%, respectively, indicating that the genetic diversity of D. sinensis was higher than that of D. dyeriana. Analysis, based on similarity coefficients, Shannon diversity index and Nei gene diversity index, also confirmed this result. AMOVA analysis demonstrated that the genetic variation of D. sinensis within and among populations accounted for 56.89% and 43.11% of the total variation, respectively, and that of D. dyeriana was 57.86% and 42.14%, respectively. The Shannon diversity index and Nei gene diversity index showed similar results. The abovementioned characteristics indicated that the genetic diversity levels of these two species were extremely similar and that the interpopulational genetic differentiation within both species was relatively high. Analysis of the genetic distance among populations also supported this conclusion. Low levels of interpopulational gene flow within both species were believed to be among the leading causes for the above-mentioned phenomenon. The correlation analysis between genetic and geographical distances showed the existence of a remarkably significant correlation between the genetic distance and the longitudinal difference among populations of D. sinensis (p < 0.01), while no significant correlation was found between genetic and geographical distances among populations of D. dyeriana. This indicated that genetic distance was correlated with geographical distances on a large scale rather than on a small scale. This result may be related to differences in the selection pressure on species by their habitats with different distribution ranges. We suggest that in situ conservation efforts should focus on establishing more sites to protect the natural populations and their habitats. Ex situ conservation efforts should focus on enhancing the exchange of seeds and seedlings among populations to facilitate gene exchange and recombination, and to help conserve genetic diversity. __________ Translated from Acta Phytoecologica Sinica, 2005, 29(5): 785–792 [译自: 植物生态学报, 2005, 29(5): 785–792]     

High dynamic range characterization of the trauma patient plasma proteome

Although human plasma represents an attractive sample for disease biomarker discovery, the extreme complexity and large dynamic range in protein concentrations present significant challenges for characterization, candidate biomarker discovery, and validation. Herein we describe a strategy that combines immunoaffinity subtraction and subsequent chemical fractionation based on cysteinyl peptide and N-glycopeptide captures with two-dimensional LC-MS/MS to increase the dynamic range of analysis for plasma. Application of this "divide-and-conquer" strategy to trauma patient plasma significantly improved the overall dynamic range of detection and resulted in confident identification of 22,267 unique peptides from four different peptide populations (cysteinyl peptides, non-cysteinyl peptides, N-glycopeptides, and non-glycopeptides) that covered 3,654 different proteins with 1,494 proteins identified by multiple peptides. Numerous low abundance proteins were identified, exemplified by 78 "classic" cytokines and cytokine receptors and by 136 human cell differentiation molecules. Additionally a total of 2,910 different N-glycopeptides that correspond to 662 N-glycoproteins and 1,553 N-glycosylation sites were identified. A panel of the proteins identified in this study is known to be involved in inflammation and immune responses. This study established an extensive reference protein database for trauma patients that provides a foundation for future high throughput quantitative plasma proteomic studies designed to elucidate the mechanisms that underlie systemic inflammatory responses.     

Normalization approaches for removing systematic biases associated with mass spectrometry and label-free proteomics

Central tendency, linear regression, locally weighted regression, and quantile techniques were investigated for normalization of peptide abundance measurements obtained from high-throughput liquid chromatography-Fourier transform ion cyclotron resonance mass spectrometry (LC-FTICR MS). Arbitrary abundances of peptides were obtained from three sample sets, including a standard protein sample, two Deinococcus radiodurans samples taken from different growth phases, and two mouse striatum samples from control and methamphetamine-stressed mice (strain C57BL/6). The selected normalization techniques were evaluated in both the absence and presence of biological variability by estimating extraneous variability prior to and following normalization. Prior to normalization, replicate runs from each sample set were observed to be statistically different, while following normalization replicate runs were no longer statistically different. Although all techniques reduced systematic bias to some degree, assigned ranks among the techniques revealed that for most LC-FTICR-MS analyses linear regression normalization ranked either first or second. However, the lack of a definitive trend among the techniques suggested the need for additional investigation into adapting normalization approaches for label-free proteomics. Nevertheless, this study serves as an important step for evaluating approaches that address systematic biases related to relative quantification and label-free proteomics.     

Generation and Characterization of Thymidine/d-Alanine Auxotrophic Recombinant Lactococcus lactis subsp. lactis IL1403 Expressing BmpB

Genetic engineering of Lactococcus lactis to produce a heterologous protein may cause potential risks to the environment despite the industrial usefulness of engineered strains. To reduce the risks, we generated three auxotrophic recombinant L. lactis subsp. lactis IL1403 strains expressing a heterologous protein, BmpB, using thyA- and alr-targeting integration vectors: ITD (thyA ⁻ alr ⁺ bmpB ⁺), IAD (thyA ⁺ alr ⁻ bmpB ⁺), and ITDAD (thyA ⁻ alr ⁻ bmpB ⁺). After construction of integration vectors, each vector was introduced into IL1403 genome. Integration of BmpB expression cassette, deletion of thyA, and inactivation of alr were verified by using PCR reaction. All heterologous DNA fragments except bmpB were eliminated from those recombinants during double crossover events. By using five selective agar plates, we also showed thymidine auxotrophy of ITD and ITDAD and d-alanine auxotrophy of IAD and ITDAD. In M17G and skim milk (SYG) media, the growth of the three recombinants was limited. In MRS media, the growth of IAD and ITDAD was limited, but ITD showed a normal growth pattern as compared with the wild-type strain (WT). All the recombinants showed maximal BmpB expression at an early stationary phase when they were cultivated in M17G supplemented with thymidine and d-alanine. These results suggest that auxotrophic recombinant L. lactis expressing a heterologous protein could be generated to reduce the ecological risks of a recombinant L. lactis.     

Overexpression of SERCA2a by gene transfer enhances myocardial systolic function in canines

The present study is aimed to study the effect of sarcoplasmic reticulum Ca(2+)-ATPase 2a (SERCA2a) gene transfer on the contractile function of isolated cardiomyocytes of canines. The cardiomyocytes were isolated with collagenases. The isolated cardiac cells were divided into untransfected group, empty vector group and SERCA2a-transfected group. Recombinant adenovirus vector carrying enhanced green fluorescent protein gene was used for SERCA2a gene delivery. The expression of SERCA2a protein in cardiomyocytes was determined by Western blot. Contractile function of cardiomyocytes was measured with motion edge-detection system of single cell at 48 h after transfection. The results showed, compared with untransfected group, SERCA2a protein level, percentage of peak contraction amplitude under normal condition, percentages of peak contraction amplitude under Ca(2+) or isoproterenol stimulation, time-to-peak contraction (TTP) and time-to-50% relaxation (R50) in SERCA2a-transfected group all increased significantly. While all the above indices in empty vector group did not show any differences with those in untransfected group. These results suggest that the overexpression of SERCA2a by gene transfer may enhance the contraction function of canine myocardial cells.     

Dynamic monitoring the TCR CDR3 spectratypes in patients with metastatic CRC treated with a combination of bevacizumab, irinotecan, fluorouracil, and leucovorin

In the present study, either modified IFL regimen (modified irinotecan, fluorouracil and leucovorin, mIFL) alone or in combination with bevacizumab was used to treat patients with metastatic colorectal cancer (CRC). Treatment efficacy was assessed using coupled tomography imaging diagnosis. The toxicity accompany with treatment was evaluated, as well as T cell receptor (TCR) repertoire before and several cycles after therapy was dynamically monitored by analyzing the complementarity-determining region 3 (CDR3) length distribution within CD4⁺ and CD8⁺ T cell subsets. The degrees of normalization of the T cell repertoire in CRC patients treated with the two methods were compared. The results showed that mIFL combined with bevacizumab was more effective in treating patients with metastatic CRC, and was accompanied by an increase in side effects such as proteinuria and hematuria. An even more restricted CDR3 profile in patients with metastatic CRC compared with healthy control has been detected. A prominent usage of TCR β chain variable (BV) gene BV12 and BV16 families within the CD4⁺ T cell subset and BV19 and BV21 families within the CD8⁺ T cell subset have been found before treatment. Moreover, CD8⁺ T cells showed more restricted patterns than CD4⁺ T cells, especially in patients before treatment. For patients with stable disease (SD) or partial remission (PR) after treatment, a less restricted CDR3 profile in post-treatment compared with pre-treatment has been found, but the opposite result was observed for patients with progressive disease (PD). The less restricted CDR3 pattern suggested a trend toward normalization of the TCR repertoire. The normalization of TCR repertoire significantly increased in patients treated with mIFL in combination with bevacizumab, but slightly in patients treated with mIFL alone. The results demonstrate a positive correlation between post-therapy TCR repertoire normalization and remission of metastatic CRC.     

Reactivation of a silenced minimal Mutator transposable element system following low-energy nitrogen ion implantation in maize

In maize, Mutator transposable elements are either active or silenced within the genome. In response to environmental stress, silenced Mutator elements could be reactivated, leading to changes in genome structure and gene function. However, there is no direct experimental evidence linking environmental stress and Mutator transposon reactivation. Using a maize line that contains a single inactive MuDR and a lone nonautonomous Mutator element, a Mu1 insertion in the recessive reporter allele a1-mum2 in an inactive Mutator background, we directly assessed Mutator reactivation following low-energy nitrogen ion implantation. We observed that N⁺ implantation decreased cytosine methylation in MuDR terminal inverted repeats and increased expression of mudrA and mudrB. Both changes were associated with increased transpositional activity of MuDR through reactivation of the inactive minimal Mutator transposable element system. This study provides direct evidence linking environmental stress agents and Mutator transposon mobilization in maize. In addition, the observed changes to DNA methylation suggest a new mechanism for mutations by low-energy ion implantation.     

Synthesis and biological evaluation of isoflavone fatty acid esters with potential weight loss and hypolipidemic activities

A series of isoflavone fatty acid esters were designed on the basis of endogenous oleoyl-estrone using estrogen moiety modification strategy. Ten new compounds were synthesized, and their body weight loss and hypolipidemic bioactivities were assayed. Some of these novel compounds could effectively inhibit the differentiation of 3T3-L1 preadipocytes in vitro. The most potent compound 1a significantly decreased the body weight and white adipose tissue weight in a high-fat diet-induced rat model. Also, compound 1a showed good hypolipidemic activity and low toxicity.     

中国亚热带地区柑桔的气候适宜性

应用生态适宜度理论和模糊数学方法,建立柑桔气候适宜度模型,对中国亚热带地区柑桔生产的气候适宜性及其时空差异进行了计算、评价,并分析了中国亚热带地区柑桔各生育期气候适宜性以及中国亚热带地区各县区的平均气候适宜度.结果表明:柑桔花芽分化期、发芽期、果实成熟期的适宜性较低、风险较高,其他各生育期的适宜性较高、风险较低,影响中国亚热带地区柑桔生产的关键问题是冷害和伏旱.柑桔温度适宜性的地域分异呈纬度地带性规律;降水适宜度最适宜型分布在亚热带中部"射阳-那坡"一线区域,其东南为适宜型区,西北为次适宜型区,高山区为不适宜型区;日照适宜度与实际日照时数的区域变化规律基本一致,大致呈高纬度区高于低纬度区、高海拔区高于低海拔区;受温度因子的限制,气候适宜度与温度适宜度的分布情况大体一致,南部大于北部,基本也呈纬度地带性规律.从中国亚热带地区柑桔气候适宜度的年际变化分析可知,整个亚热带地区柑桔气候适宜度呈下降趋势,且存在区域差异,说明气候变化引起了中国亚热带柑桔适宜区和关键生育期的变动.     

在脉冲污染环境中具有阶段结构的捕食食饵Gompertz模型的动力学性质

研究了在脉冲污染环境中,捕食者具有阶段结构的捕食食饵Gompertz模型的动力学性质,获得了捕食者灭绝周期解全局吸引和系统持续生存的条件.