DNA degradation is a biochemical hallmark in apoptosis. It has been demonstrated in many cell types that there are two stages of DNA fragmentation during the apoptotic execution. In the early stage, chromatin DNA is c...DNA degradation is a biochemical hallmark in apoptosis. It has been demonstrated in many cell types that there are two stages of DNA fragmentation during the apoptotic execution. In the early stage, chromatin DNA is cut into large molecular weight DNA fragments, although the responsible nuclease(s) has not been recognized. In the late stage, the chromatin DNA is cleaved further into short oligonucleosomal fragments by a well-characterized nuclease in apoptosis,the caspase-activated DNase (CAD/DFF40). In this study, we demonstrate that large molecular weight DNA fragmentation also occurs in Xenopus egg extracts in apoptosis. We show that the large molecular weight DNA fragmentation factor (LDFF) is not the Xenopus CAD homolog XCAD. LDFF is activated by caspase-3. The large molecular weight DNA fragmentation activity of LDFF is Mg2+-dependent and Ca2+-independent, can occur in both acidic and neutral pH conditions and can tolerate 45℃ treatment. These results indicate that LDFF in Xenopus egg extracts might be a new DNase (or DNases) responsible for the large DNA fragmentation.展开更多
Synthetic biology is a newly developed field of research focused on designing and rebuilding novel biomolecular components, circuits, and networks. Synthetic biology can also help understand biological principles and ...Synthetic biology is a newly developed field of research focused on designing and rebuilding novel biomolecular components, circuits, and networks. Synthetic biology can also help understand biological principles and engineer complex artificial metabolic systems. DNA manipulation on a large genome-wide scale is an inevitable challenge, but a necessary tool for synthetic biology. To improve the methods used for the synthesis of long DNA fragments, here we constructed a novel shuttle vector named p GF(plasmid Genome Fast) for DNA assembly in vivo. The BAC plasmid p CC1 BAC, which can accommodate large DNA molecules, was chosen as the backbone. The sequence of the yeast artificial chromosome(YAC) regulatory element CEN6-ARS4 was synthesized and inserted into the plasmid to enable it to replicate in yeast. The selection sequence HIS3, obtained by polymerase chain reaction(PCR) from the plasmid p BS313, was inserted for screening. This new synthetic shuttle vector can mediate the transformation-associated recombination(TAR) assembly of large DNA fragments in yeast, and the assembled products can be transformed into Escherichia coli for further amplification. We also conducted in vivo DNA assembly using p GF and yeast homologous recombination and constructed a 31-kb long DNA sequence from the cyanophage PP genome. Our findings show that this novel shuttle vector would be a useful tool for efficient genome-scale DNA reconstruction.展开更多
The maximal matching problem （MMP） is to find maximal edge subsets in a given undirected graph, that no pair of edges are adjacent in the subsets. It is a vitally important NP-complete problem in graph theory and ap...The maximal matching problem （MMP） is to find maximal edge subsets in a given undirected graph, that no pair of edges are adjacent in the subsets. It is a vitally important NP-complete problem in graph theory and applied mathematics, having numerous real life applications in optimal combination and linear programming fields. It can be difficultly solved by the electronic computer in exponential level time. Meanwhile in previous studies deoxyribonucleic acid （DNA） molecular operations usually were used to solve NP-complete continuous path search problems, e.g. HPP, traveling salesman problem, rarely for NP-hard problems with discrete vertices or edges solutions, such as the minimum vertex cover problem, graph coloring problem and so on. In this paper, we present a DNA algorithm for solving the MMP with DNA molecular operations. For an undirected graph with n vertices and m edges, we reasonably design fixed length DNA strands representing vertices and edges of the graph, take appropriate steps and get the solutions of the MMP in proper length range using O（n^3） time. We extend the application of DNA molecular operations and simultaneously simplify the complexity of the computation.展开更多
基金This work was supported by Major State Basic Research Program of China(No.G1999053905)National Science Fund for Distinguished Young Scholars(No.30225016).
文摘DNA degradation is a biochemical hallmark in apoptosis. It has been demonstrated in many cell types that there are two stages of DNA fragmentation during the apoptotic execution. In the early stage, chromatin DNA is cut into large molecular weight DNA fragments, although the responsible nuclease(s) has not been recognized. In the late stage, the chromatin DNA is cleaved further into short oligonucleosomal fragments by a well-characterized nuclease in apoptosis,the caspase-activated DNase (CAD/DFF40). In this study, we demonstrate that large molecular weight DNA fragmentation also occurs in Xenopus egg extracts in apoptosis. We show that the large molecular weight DNA fragmentation factor (LDFF) is not the Xenopus CAD homolog XCAD. LDFF is activated by caspase-3. The large molecular weight DNA fragmentation activity of LDFF is Mg2+-dependent and Ca2+-independent, can occur in both acidic and neutral pH conditions and can tolerate 45℃ treatment. These results indicate that LDFF in Xenopus egg extracts might be a new DNase (or DNases) responsible for the large DNA fragmentation.
基金supported by the 973 program,Grant No.2012CB721102
文摘Synthetic biology is a newly developed field of research focused on designing and rebuilding novel biomolecular components, circuits, and networks. Synthetic biology can also help understand biological principles and engineer complex artificial metabolic systems. DNA manipulation on a large genome-wide scale is an inevitable challenge, but a necessary tool for synthetic biology. To improve the methods used for the synthesis of long DNA fragments, here we constructed a novel shuttle vector named p GF(plasmid Genome Fast) for DNA assembly in vivo. The BAC plasmid p CC1 BAC, which can accommodate large DNA molecules, was chosen as the backbone. The sequence of the yeast artificial chromosome(YAC) regulatory element CEN6-ARS4 was synthesized and inserted into the plasmid to enable it to replicate in yeast. The selection sequence HIS3, obtained by polymerase chain reaction(PCR) from the plasmid p BS313, was inserted for screening. This new synthetic shuttle vector can mediate the transformation-associated recombination(TAR) assembly of large DNA fragments in yeast, and the assembled products can be transformed into Escherichia coli for further amplification. We also conducted in vivo DNA assembly using p GF and yeast homologous recombination and constructed a 31-kb long DNA sequence from the cyanophage PP genome. Our findings show that this novel shuttle vector would be a useful tool for efficient genome-scale DNA reconstruction.
文摘The maximal matching problem （MMP） is to find maximal edge subsets in a given undirected graph, that no pair of edges are adjacent in the subsets. It is a vitally important NP-complete problem in graph theory and applied mathematics, having numerous real life applications in optimal combination and linear programming fields. It can be difficultly solved by the electronic computer in exponential level time. Meanwhile in previous studies deoxyribonucleic acid （DNA） molecular operations usually were used to solve NP-complete continuous path search problems, e.g. HPP, traveling salesman problem, rarely for NP-hard problems with discrete vertices or edges solutions, such as the minimum vertex cover problem, graph coloring problem and so on. In this paper, we present a DNA algorithm for solving the MMP with DNA molecular operations. For an undirected graph with n vertices and m edges, we reasonably design fixed length DNA strands representing vertices and edges of the graph, take appropriate steps and get the solutions of the MMP in proper length range using O（n^3） time. We extend the application of DNA molecular operations and simultaneously simplify the complexity of the computation.
