细胞功能测定

The polymerase chain reaction (PCR) (1,2) is now a fundamental tool of molecular biology. Although PCR provides the basis for a variety of sensitive analytical techniques, it can also be used in a synthetic capacity to generate large quantities of specific DNA fragments. The alteration of ampli ...

Protocols for site-directed mutagenesis are widely used in molecular biology and include many polymerase chain reaction (PCR)-based methods that have been developed in order to achieve efficient mutagenesis of a target DNA sequence (1–4). However, some of these methods described req ...

Before the advent of polymerase chain reaction (PCR) technology, many methods for site-directed mutagenesis basically relied on enzymatic extension of a mutagenic oligonucleotide annealed to a single-stranded template and amplification of the ligase-sealed double-stran ...

An important manipulation in molecular genetics is to make ordered deletions into a cloned piece of DNA. The most widely used application of this method is in DNA sequencing. Ordered deletions can also be used in delineating sequences that are important for the function of a gene, such as those requi ...

In vitro site-directed mutagenesis has been widely used in vector modification, and in gene and protein structure/function studies (1,2). This procedure typically employs one or more oligonuleotides to introduce defined mutations into a DNA target of known sequence (2–9). A variation of ...

The generation of nested deletions within cloned fragments of DNA has important applications in molecular biology. For DNA sequencing, nested deletions provide a series of overlapping templates that can be used to generate a composite sequence with a single sequencing primer; in gene and ...

Oligonucleotides can bind in the major groove of duplex DNA and form triple helices in a sequence-specific manner 1–4). Progress in elucidating the third strand binding code has raised the possibility of developing nucleic acids as sequence-specific reagents for research and possibly c ...

A number protocols have been established for site-directed mutagenesis based on the work of Smith (1) and Hutchinson et al. (2) that use hybridization of a mismatched oligonucleotide to a DNA template followed by second-strand synthesis by a DNA polymerase. These techniques provide effici ...

For convenience the term quiescent cells will be used in this chapter to define normal or tumor cells, untreated by any specific drugs, that do not progress through the mitotic cycle for an extended period of time. The definition is purposefully wide to cover a variety of situations, often with differ ...

Analysis of the cell cycle was significantly advanced by the development of photometric methods for quantitative measurement of biochemical constituents in single cells. Population biochemical analysis could then be performed on a cell-by-cell basis, and distinct subpopulat ...

The dispersal of solid tissues into a single-cell suspension has become an integral component of many current techniques for quantitative cytokinetic analysis of both normal and neoplastic tissues. This is particularly evident in the application of flow cytometric (FCM) procedur ...

The measurement of percent labeled mitoses (PLM, or alternatively, FLM, for fraction of labeled mitoses) was developed in 1959 by Quastler and Sherman (8) for estimating the durations of the component phases of the cell cycle.

相关专题 试想一下，若没有各种方法来分离组成细胞的不同区室和组分，那么现代生物学将困难重重。从细胞信号通路到新陈代谢，人们一直想知道，它们在哪儿，过去、现在及将来。 赛默飞世尔科技的市场部经理Monica O’Hara Noonan介绍，研究人员对细胞进行分级分离(fractionate)的原因有很多。他们可能希望，当感兴趣的蛋白在某些条件下或某种处理之后转位时，对其进行追踪。对细胞进行分级分离能增强低丰度蛋白的检测能力，或降低 ...

A combination of forward and reverse genetic approaches in zebrafish has revealed novel roles for canonical Wnt and Wnt/PCP signaling during vertebrate development. Forward genetics in zebrafish provides an exceptionally powerful tool to assign roles in vertebrate developmen ...

The Wnt pathway is a highly conserved signal transduction pathway that plays many critical roles in early animal development. Recent studies have shown that this pathway plays a conserved role in the specification and patterning of the animal – vegetal (A – V) axis in sea urchins and sea anemones. Th ...

The animal–vegetal (A–V) axis is a maternally established asymmetry that is present in most animal eggs, and it plays an important role in germ-layer segregation. Recent work has shown that the canonical Wnt signaling pathway plays an evolutionarily conserved role in specifying and patter ...

Wnt signaling regulates a remarkably diverse array of cellular and developmental events during animal embryogenesis and homeostasis. The crucial role that Wnt signaling plays in regulating axial patterning in early embryos has been particularly striking. Recent work has highli ...

Genetic and biochemical analyses in model systems such as the fruitfly, Drosophila melanogaster, have successfully identified several genes that play key regulatory roles in fundamental cellular and developmental processes. However, the analyses of the complete genome seque ...

The secreted morphogen Wingless (Wg) has a variety of functions throughout Drosophila eye development, controlling tissue specification, growth, and patterning. Wg plays a critical role in subdividing the eye imaginal disc into separate primordia that will give rise to the adult reti ...

“ Tissue ” or “ planar ” polarity is a characteristic of many epithelial tissues and is not only required for proper cell alignment, but in many instances is absolutely essential for normal function. Planar cell polarity (PCP) is the polarization of cells within the plane of an epithelium in a direction ...