Twostage Model Of Recombination

For recombination to occur, homologous chromosomes must pair up with their DNA sequences aligned. Studies in yeast have led to a rather complex model for this process. The complications occur because DNA sequence similarities or identities occur not only between the pairs of homologs but also between different chromosomes as a result of dispersed repeated DNA sequences (see Chapter 14) or dispersed gene families with similar or identical members. The recombination apparatus in bacteria, yeast,...

Bacterial colonies

And release bacteriophage, then a convenient method of cloning is to grow a continuous layer of susceptible bacteria and then cover the culture with a thin layer of dilute bacteriophage suspension. Individual bacteriophage will infect cells, leading to continual cycles of bacteriophage growth and cell lysis until visible holes in the bacterial lawn, called bacteriophage plaques, are observed. Individual colonies or plaques are selected for further study by picking, literally by sampling with a...

Detection Of Homozygous Regions

Because the human species is highly outbred, homozygous regions are rare. Such regions can be found, however, by traditional methods or by some of the fairly novel methods that will be described in Chapter 13. Homozygous regions are very useful both in the diagnosis of cancer and in certain types of genetic mapping. The significance of homozygous regions in cancer is shown in Figure 6.37. Oncogenes are ordinary cellular genes, or foreign genes that under appropriate circumstances can lead to...

Good amplification after first few cycles

Figure 4.6 Introducing extra functionalities by appropriately designed primers. (a) Incorporation of a 5'-terminal label (asterisk). (b) Incorporation of flanking restriction sites, useful for subsequent cloning. (c) Compensating for less than optimal length initial sequence information. This problem typically arises when one is trying to walk by from a known region of the genome into flanking unknown regions. Then one starts with a bit of known sequence at the extreme edge of the charted...

Thermodynamics Of Imperfectly Paired Duplexes

In contrast to the small number of discrete interactions that must be considered in calculating the energetics of perfectly paired DNA duplexes, there is a plethora of ways that duplexes can pair imperfectly. We have available model compound data on most of these so that estimates of the energetics can be made. However, the large number of possibilities precludes a complete analysis of imperfections in the context of all possible sequences, at least for the present. The simplest imperfection is...

Subchromosomal Mapping Panels

Frequently it is possible to assemble sets of cell lines containing fragments of only one chromosome of interest. The occurrences of a DNA probe sequence in a subset of these cell lines can be used to assign the probe to a specific region of the chromosome. The accuracy of that assignment is determined by the accuracy with which the particular chromosome fragments contained in each cell line are known. This can vary quite considerably, but in general this approach is quite a powerful and...

Basic Principles Of The Polymerase Chain Reaction

What makes PCR a tool of immense power and flexibility is the requirement of DNA polymerases for pre-existing DNA primers. Thus DNA polymerases cannot start DNA chains de novo a primer can be used to determine where, along a DNA template, the synthesis of the complement of that stand begins. It is this primer requirement that allows the selective amplification of any DNA region by using appropriate, specific DNA primers. Once started, a DNA polymerase like E. coli DNA polymerase I pol I will...

Double Helical Structure

The two common base pairs A-T and G-C are well-known, and little evidence for other base interactions within the DNA's double helix exists. The key feature of double-helical DNAs duplexes , which dominates their properties, is that an axis of symmetry relates the two strands Box 1.1 . Figure 1.1 Structure of the phosphodiester backbone of DNA and RNA, and the four major bases found in DNA and RNA the purines, adenine A and guanine G , and the pyrimidines, cytosine C and thymine T or uracil U ....

C2 Symmetry

Base Pair Per Helical Turn

C2 symmetry implies that a structure is composed of two identical parts. An axis of rotation can be found that interchanges these two parts by a 180-degree rotation. This axis is called a C2 axis. An example of a common object that has such symmetry is the stitching of a baseball, which is used to bring two figure-8 shaped structures together to make a spherical shell. An example of a well-known biological structure with C2 symmetry is the hemoglobin tetramer which is made up of two identical...

Pcr Noise Mispriming

Typical PCR conditions with two convergent primers offer a number of possible unintended primed amplifications. These are illustrated in Figure 4.4. If the primers are not chosen wisely, one of the two primers may be able to act alone to amplify DNA as shown in Figure 4.4b. Alternatively, the two convergent primers may have more than one site in the target that allows amplification. There is no way to plan for these events, unless the entire sequence of the sample is known. However, the chances...

Unusual Characteristics Of Sex Chromosomes And Mitochondria

In mammals a female carries two copies of the X chromosome males have one X and one Y. However, this simple difference in karyotype the set of chromosomes has profound effects that go beyond just the establishment of sex. The first thing to consider is why we need sex at all. In species with just one sex, each organism can reproduce clon-ally. The offspring of that organism may be identical. If the organism inhabits a wide ecological range, different selection processes will produce a...

Physical Mapping

WHY HIGH-RESOLUTION PHYSICAL MAPS ARE NEEDED Physical maps are needed because ordinary human genetic maps are not detailed enough to allow the DNA that corresponds to particular genes to be isolated efficiently. Physical maps are also needed as the source for the DNA samples that can serve as the actual substrate for large-scale DNA sequencing projects. Genetic linkage mapping provides a set of ordered markers. In experimental organisms this set can be almost as dense as one wishes. In humans...

Quantitative Highresolution Fish

Fiber Fish With Gene Specific Probe

The quantitative analysis of distances in high-resolution FISH has been pioneered by two groups headed by Barb Trask, currently at the University of Washington, and Jeanne Lawrence, at the University of Massachusetts in Amherst. Others have learned their methods and begun to practice them. A few representative analyses are shown in Figures 7.18 and 7.19. The distribution of measured distances between two fixed markers in individual samples of interphase chromatin varies over quite a wide range,...

Padlock Probes

Ulf Landegren and his collaborators recently demonstrated that DNA topology can be used to improve the stringency of detection of correctly hybridized DNA probes. A schematic view of the procedure that they have developed is shown in Figure 3.20. Consider the hybridization of a linear single-stranded probe to a single-stranded circular target. The probe is specially designed so that it's 3'- and 5'-terminal sequences are complementary to a continuous block of DNA sequence in the target. Probes...