Ligation-assisted homologous recombination enables precise genome editing by deploying both MMEJ and HDR

CRISPR/Cas12a is a single effector nuclease that, like CRISPR/Cas9, has been harnessed for genome editing based on its ability to generate targeted DNA double strand breaks (DSBs). Unlike the blunt-ended DSB generated by Cas9, Cas12a generates sticky-ended DSB that could potentially aid precise genome editing, but this unique feature has thus far been underutilized. In the current study, we found that a short double-stranded DNA (dsDNA) repair template containing a sticky end that matched one of the Cas12a-generated DSB ends and a homologous arm sharing homology with the genomic region adjacent to the other end of the DSB enabled precise repair of the DSB and introduced a desired nucleotide substitution. We termed this strategy ‘Ligation-Assisted Homologous Recombination’ (LAHR). Compared to the single-stranded oligo deoxyribonucleotide (ssODN)-mediated homology directed repair (HDR), LAHR yields relatively high editing efficiency as demonstrated for both a reporter gene and endogenous genes. We found that both HDR and microhomology-mediated end joining (MMEJ) mechanisms are involved in the LAHR process. Our LAHR genome editing strategy, extends the repertoire of genome editing technologies and provides a broader understanding of the type and role of DNA repair mechanisms involved in genome editing.     

宁夏黄河流域景观破碎化时空变化特征

基于RS和GIS技术,以土地利用数据为基础,采用有效粒度尺寸对宁夏黄河流域1985年和2010年的景观破碎化进行时空变化分析。结果显示:流域整体的有效粒度尺寸(meff)从1985年的6326.62 km2下降到2010年的2974.32 km2,破碎化程度呈显著加剧变化;从流域内部来看,景观破碎化程度最大的是黄左区间,破碎化程度最小的是苦水河和红柳沟;除引黄区间外,其余分区景观破碎化程度在25年间均有所加剧。特征尺度分析结果显示研究区景观破碎化空间变异分析的合适尺度为4500 m;景观破碎化指数的空间变异结果表明破碎化程度较大的区域面积明显多于破碎化程度弱的区域,景观破碎化空间异质性在25年间表现出明显的上升趋势。在海拔背景条件下,2400 m以下区域的景观破碎化程度较高,2400m以上区域破碎化程度较低,且随海拔的升高有降低的趋势;流域景观破碎化受人为干扰影响强烈,在人为干扰较强的1000—1500 m区段的景观破碎化程度最大;景观破碎化在时间上的变化受人为干扰影响产生的变化最为显著,由自然条件改变产生的影响有限。研究结果可为西北地区景观格局及景观破碎化的研究提供参考,并为区域景观格局优化和土地的有效管理提供依据。     

岛屿生物地理学与集合种群理论的本质与渊源

岛屿生物地理学和集合种群理论是目前生物多样性保育所依赖的主要生态学理论。人们通常强调这两种理论的区别,对它们之间的关联却很少注意到。事实上,这两种理论是同根同源的。以经典集合种群理论的创始者R．Levim对他与岛屿生物地理学的创始者R．H．MaeArthur的合作过程以及岛屿生物地理学对他提出集合种群理论的影响的回顾为基础,分析比较了岛屿生物地理学、经典集合种群理论、以Hanski为代表的现代集合种群理论的基本假设、研究范式和核心思想的异同,简要介绍了多物种集合种群与集合群落研究的差异,最后分析了岛屿生物地理学和集合种群理论在生物多样性保育实践中的应用和存在问题。     

捕食者-猎物关系的理论和应用研究

捕食者和猎物相互关系的研究,长期以来一直是动物生态学研究的中心课题之一。这种研究可区分为3种类型:第一是理论研究,即组建各种数学模型以便模拟捕食者和猎物间的相互关系;第二是实验种群研究,通常是在实验室内选用原生动物和节肢动物实验种群对捕食一猎物间的动态关系进行观察,并将观察结果与理论模型进行比较;第三是在田间对自然种群中的捕食者-猎物关系进行研究,并利用从理论研究和实验种群研究中所总结出来的各种基本原理对观察资料进行分析。在应用生态学领域中,常常靠引进新的更加有效的天敌来控制各种害虫。这些实际工作有些已获得成功,也有不少未取得预期效果,不管成功与否,它们都可被看作是对捕食者-猎物关系所进行的田间实验.     

Association between polymorphisms in the promoter region of interleukin-10 and susceptibility to inflammatory bowel disease

The aim of this study was to assess the association of polymorphisms in the promoter region of the IL-10 gene with the risk of inflammatory bowel disease (IBD), including Crohn’s disease (CD) and ulcerative colitis (UC). Fifteen studies (3,693 cases and 4,574 controls) were included in a meta-analysis of association between IL-10 ?1082G/A, ?819C/T and ?592C/A polymorphisms, and IBD, CD and UC using allele contrast and the recessive, dominant, and additive models. Hardy–Weinberg equilibrium was confirmed for each study. Heterogeneity and study quality were investigated using stratification analyses and sensitivity analyses. Polymorphism ?1082G/A showed significant association with CD, with odds ratios (ORs) for the GG + GA genotype and GG versus AA genotype of 1.278 (1.004–1.627) and 1.238 (1.027–1.492) in all subjects. Significant associations were found in the Caucasian subgroup using the allele contrast, dominant, and additive models. C-allele carriers of the ?819C/T polymorphism were at increased risk of IBD (OR 1.093, 95 % CI 1.004–1.190). Association with the ?819C/T polymorphism was also found in Caucasians with CD (C vs. T: OR 1.104, 95 % CI 1.010–1.206; CC + CT vs. TT: OR 1.328, 95 % CI 1.006–1.754; CC vs. TT: OR 1.339, 95 % CI 1.008–1.778), and with UC (CC vs. CT + TT: OR 1.188, 95 % CI 1.019–1.385). No significant association was found between the ?592C/A polymorphism and IBD, CD or UC. In conclusion, the meta-analysis demonstrated clear association between the IL-10 polymorphisms ?1082G/A and ?819C/T and the risk of IBD.     

长白山阔叶红松林凋落物组成及其季节动态

为了解群落尺度上凋落物组成及其时空变化,对长白山阔叶红松林25 hm² 样地内2008年度收集的凋落物进行统计分析.结果表明: 一年间,150个收集器内收集的凋落叶分属35种树木,占样地内胸径≥1 cm树种数(52种)的67.3%;凋落物总量为29.39 kg,折合3918.4 kg·hm^-2,其中,阔叶、杂物、针叶和枝条凋落量分别占凋落物总量的61.7%、18.0%、11.7%和 8.6%;紫椴、水曲柳、蒙古栎、色木槭和春榆5个树种的叶凋落量占阔叶总凋落量的83.8%;不同树种的凋落量季节差异很大,61.9%的凋落物产生于9月13日至10月10日.其中,红松和紫椴叶凋落高峰出现在9月13-26日,蒙古栎、春榆和色木槭叶凋落高峰出现在9月27日-10月10日.收集器间凋落物量差异较大,其中68个收集器的年凋落量在150～200 g,1个收集器大于500 g;单个收集器全年最多可收集到18个树种的凋落叶,凋落叶种数为12种的收集器最多(32个).叶凋落量与样地内该树种的胸高断面积总和成正比.样地内凋落叶的分布存在明显的空间异质性,且收集器内凋落物的收集量与其所处的位置有关.     

In vitro insulin refolding: Characterization of the intermediates and the putative folding pathway

The in vitro refolding process of the double-chain insulin was studied based on the investigation of in vitro single-chain insulin refolding. Six major folding intermediates, named P1A, P2B, P3A, P4B, P5B, and P6B, were captured during the folding process. The refolding experiments indicate that all of these intermediates are on-pathway. Based on these intermediates and the formation of hypothetic transients, we propose a two-stage folding pathway of insulin. (1) At the early stage of the folding process, the reduced A chain and B chain individually formed the intermediates two A chain intermediates (P1A and P3A), and four B chain intermediates (P2B, P4B, P5B, and P6B). (2) In the subsequent folding process, transient Ⅰ was formed from P3A through thiol/disulfide exchange reaction; then, transients Ⅱ and Ⅲ, each containing two native disulfides, were formed through the recognition and interaction of transient Ⅰ with P4B or P6B and the thiol group's oxidation reaction mainly using GSSG as oxidative reagent; finally, transients Ⅱ and Ⅲ, through thiol/mixture disulfide exchange reaction, formed the third native disulfide of insulin to complete the folding.     

单克隆抗体γ3重链可变区cDNA的合成与克隆

用异硫氰酸胍法从分泌单克隆抗体的杂交瘤细胞中提取总RNA,经oligo(dT)-纤维素柱亲和层析获得poly(A)~ RNA后,用恒定区5′端第122—125号氨基酸密码的互补序列3′A-T-A-G-G-T-G-A-C-C 5′做为引物,进行逆转录酶反应,合成双链cDNA,大小为300bp左右,与重链可变区基因的长度相符。用dC:dG接尾的方法,将ds-cDNA插入pUC19质粒,转化E.coli HB101。分离出重组体之后,经菌落原位杂交,酶切重组质粒DNA及Southern印迹,证明插入片段是重链可变区基因。     

围隔生态系内浮游植物对富磷的响应

在长江口外应用围隔生态系实验方法,研究了富磷与浮游植物生物量的相关性以及浮游植物对磷吸收的种间竞争。结果表明,可溶性磷(主要是PO     

海洋生态资本价值结构要素与评估指标体系

针对海洋生态系统的特点,构建了海洋生态资本价值的结构要素及其评估指标体系。海洋生态资本的价值是指海洋生态资本的存量价值及其产生的收益流价值,包括各类海洋生态资源的现存量价值及其组成海洋生态系统整体而产生的生态系统服务价值。其中,海洋生态资源的现存量价值由海洋生物资源存量价值和海洋生境资源存量价值构成。海洋生态系统服务价值包含海洋供给服务、海洋调节服务、海洋文化服务和海洋支持服务等4个子要素的价值。