一个RNAi载体上反向重复片段的测序策略

相邻的反向重复DNA片段有形成单链内二级结构的倾向,属于一种测序困难的DNA模板。解决RNAi载体插入的反向重复片段的测序问题,为该类载体正确性的测序验证奠定基础。采用常规分子克隆方法构建表达小麦TaATG2串联反向重复片段的RNAi载体,设计2种策略对经菌落PCR初步鉴定的载体进行测序验证:一种是以完整的载体质粒为模板进行测序;另一种是先对载体进行酶切处理,切除反向重复片段中的一个后对保留另一个片段的线性载体进行测序。结果表明,第一种测序策略受到串联反向重复片段形成的单链内部二级结构的影响,测序信号在反向重复片段处出现衰减或乱峰,无法读取序列。第二种测序策略排除了2个反向重复片段之间的干扰,保留在载体上的片段测序信号清晰,序列准确。采用酶切切除一个片段后进行测序的方法,经过2次酶切和2次测序可以有效地对载体上的2个反向重复片段分别进行序列测定,进而确认构建载体的正确性。     

Molecular and immunological characterization of the major outer membrane proteins of Brucella

Abstract Saccharomyces cerevisiae exponentially growing in basic or 0.7 M NaCl medium were isotopically labelled with ³⁵S-methionine, followed by protein separation and quantification by two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) combined with computerised image analysis. The electrophoretic separation resolved about 650 proteins of which 13 displayed significant and at least 2-fold changes in rate of synthesis during saline growth. By sequencing of 2D-PAGE resolved proteins, one of the 8 induced spot, p42.9/5.5, was shown to correspond to the full length (containing the N-terminal extension) product of the GPD 1 gene encoding the cytoplasmic glycerol 3-phosphate dehydrogenase. The expression of the TDH 3 gene, glyceraldehyde 3-phosphate dehydrogenase, and the ENO 2 gene, enolase, decreased during growth in NaCl medium, declines hypothesised to have an impact on the flux to glycerol.     

Isolation and characterization of polymorphic microsatellite loci in Indian major carp,Catla catla using next-generation sequencing platform

Catla catla, the second most important Indian major carp, is gaining its popularity among Indian fish farmers due to its high growth rate and consumer preferences. Simple sequence repeats (SSRs) are rapidly evolving, versatile, co-dominant and highly informative molecular markers used in genetic research. However, the time and cost involved in developing such resources has limited their extensive use. Advent of massive parallel sequencing technology has considerably eased these limitations. In the present investigation, we used Ion Torrent sequencing platform to identify potentially amplifiable microsatellite loci for catla. A modest sequencing volume generated approximately 5.7 MB of sequence data. Out of 29,794 sequences generated, 21,477 contained simple sequence repeats. Only 81 sequences had enough flanking sequences for primer designing. Out of 81 loci, 51 were successfully PCR amplified in a panel of five unrelated individuals. Out of 15 loci randomly checked for polymorphism, 13 loci were polymorphic with allele number ranged from 3 to 6 and two loci were found to be monomorphic. The observed and expected heterozygosities ranged from 0.565 to 0.870 and 0.483–0.804, respectively. These markers will be useful for studying genetics of wild populations, breeding programs of C. catla and closely related species.     

Single-nucleus sequencing deciphers developmental trajectories in rice pistils

1. Download : Download high-res image (238KB)
2. Download : Download full-size image

     

Genomic characterization of metastatic patterns from prospective clinical sequencing of 25,000 patients

1. Download : Download high-res image (241KB)
2. Download : Download full-size image

     

Tropical terrestrial invertebrates—Where to from here?

There are over one million described invertebrate species on Earth, the majority of which are likely to inhabit the highly biodiverse rain forests around the equator. These are some of the most vulnerable ecosystems on Earth due to the pressures of deforestation and climate change with many of their inhabitants at risk of extinction. Invertebrates play a major role in ecosystem functioning from decomposition and nutrient cycling to herbivory and pollination; however, while our understanding of these roles is improving, we are far from being able to predict the consequences of further deforestation, climate change, and biodiversity loss due to the lack of comparative data and the high proportion of species which remain to be discovered. As we move into an era of increased pressure on old-growth habitats and biodiversity, it is imperative that we understand how changes to invertebrate communities, and the extinction of species, affect ecosystems. Innovative and comprehensive methods that approach these issues are needed. Here, we highlight priorities for future tropical terrestrial invertebrate research such as the efficiency of sustainable land management, exploration of innovative methods for better understanding of invertebrate ecology and behavior, and quantifying the role of invertebrates in ecosystem functioning.     

The Genetic Basis of Morphological Diversity in Domesticated Goldfish

1. Download : Download high-res image (210KB)
2. Download : Download full-size image