DNA denaturation, complexity, and molecular hybridization.

the DNA
double helix structure model not only has a close relationship with its biological function, but also explains the important characteristic denaturation and complexity of DNA, which is of great significance for understanding the relationship between dna molecular structure and function.. 1. DNA denaturation (denaturation)the phenomenon of DNA molecules being loosened from stable double helix structures to irregular linear structures. The hydrogen bond that maintains double helix stability during denaturation breaks, and the accumulation force between bases is destroyed, but does not involve changes in its primary structure. Factors that can disrupt the stability of the double helix, such as
heating
, extreme pH, organic
reagents
methanol, ethanol, urea and methylamide, can cause nucleic acid molecular degeneration.
denatured DNA often under way with some changes in the nature of physiology and biology: . The viscosity of the solution is reduced. DNA double helix is a tight rigid structure, the degenerative offspring with a soft and loose irregular single linear structure, DNA viscosity has been significantly reduced.the swirl of the solution changes. After denaturation, the symmetry of the whole DNA molecule and the local structure of the molecule change, which makes the spinlight of the DNA solution change.the dechairing curve of 15-8 nucleic acidsthe hyperchromic effect. Refers to the effect of increased UV absorption of DNA solution after denaturation. The interaction of electrons between bases in DNA molecules makes DNA molecules absorb 260nm wavelengths of ultraviolet light. In the DNA double helix structure, the base is hidden inside, the DNA double helix is unziped when denaturation, so the base is exposed, and the interaction of electrons in the base is more conducive to ultraviolet absorption, so it produces a color-boosting effect.heating and denaturing double-stranded DNA, when the temperature rises to a certain height, the absorbance of the DNA solution at 260nm suddenly rises significantly to the highest value, and then the absorbance does not change significantly even as the temperature continues to rise. If the UV absorption rate of the DNA solution is graphed by temperature, the typical DNA denaturation curve is S-type.
visible DNA denaturation occurs over a very narrow temperature range. Usually during nucleic acid heating denaturation, the temperature at which the UV absorption value reaches 50% of the maximum is called the dechain temperature of the nucleic acid, because this phenomenon is similar to the melting of crystallization, also known as the melting temperature (Tm, melting temperature). At TM, 50% of the double helix structure in the nucleic acid molecule is destroyed. The Tm value of a particular nucleic acid molecule is positively related to the percentage of the total base of its G-C, and the relationship between the two can be expressed as: Tm- 69.3-0.41 (%G-C)under certain conditions (relatively short nucleic acid molecules), The size of the Tm value is also related to the length of the nucleic acid molecules, the longer the nucleic acid molecules, the greater the Tm value, in addition, when the ion strength of the solution is low, the Tm value is lower, the melting point range is wider, and vice versa, so the DNA preparation should not be stored in the solution with too low ion strength.. 2. DNA compoundingrefers to the phenomenon of denatured DNA, under appropriate conditions, in which two complementary chains are restored in whole or in part to the natural double helix structure, which is a reversal process of denaturation. Thermodynatured DNA is generally regenerative after a slow cooling, a process known as annealing. The term is also used to describe the formation of hybrid nucleic acid molecules (see below). The complexity of DNA is influenced not only by temperature, but also by other factors such as dna's own characteristics:temperature and time. It is generally believed that a temperature about 25 degrees Celsius lower than Tm is the best condition for complexity, and the farther away from this temperature, the slower the complexity speed. The temperature drop at complexity must be a slow process, and if it cools rapidly to a low temperature (e.g. below 4 degrees C) at temperatures above Tm, complexity is almost impossible, and DNA degeneration (single-stranded) status is often maintained in nucleic acid experiments in this way. This shows that the cooling time is too short and the temperature difference is not conducive to complexity.. DNA concentration. The more DNA molecules in the solution, the greater the chance of collision and binding.. The complexity of DNA sequence. The pairing of complementary bases is easier to achieve when the simple sequence of DNA molecules, such as poly (A) and poly (U), are complex. It is much more difficult to achieve complementarity in sequences with complex sequences. In nucleic acid compounding studies, a Cot term is defined (Co is the starting concentration of single-stranded DNA, t is in seconds) to represent the relationship between compounding velocity and the complexity of DNA sequence. In the discussion of the effect of DNA sequence on compound speed, the temperature, solvent ion strength, nucleic acid fragment size and other factors were fixed, and Cot was graphed with different degrees of nucleic acid molecule re-connecting part (compound rate at time t).
the non-repeated base pairs of nucleic acid molecules that show complexity above the curve. If the complexity of poly (A) is 1, the complexity of the polymer consisting of repeated (ATGC)n is 4, and the complexity of non-repeating DNA of 105 nucleotide pairs is 105. The original nucleotide
gene
groups are in a non-repetitive order, so the complexity of the non-repetitive nucleotide pairs directly reflects the size of the genome (the arrow above the figure refers to the gene size), as well as the non-repeated fragments in the genome of the ebony.
the compounding rate (called Cot1/2) measured under standard conditions (generally 0.18 ml/L cation concentration, 400 nucleotide-length fragments) is directly in direct ratio to the complexity of nucleotide pairs. For primary nucleic acid molecules, this value represents the size of the genome and the complexity of nucleotide pairs in the genome. The Resulting Cot curve is more complex because it contains many different degrees of repetitive sequences in the nuclear genome. . The principle of DNA denaturation and complexity has been widely used in medicine and life sciences. Such as nucleic acid hybridization and probe technology, polymerase chain reaction (polymerase chain reaction,
PCR
) technology. . 3. Molecular hybridization: (hybridization) nucleic acid degeneration from different sources, merged in one place for compounding, at which point, as long as the nucleotide sequence of these nucleic acid molecules contains fragments that can form complementary pairings of bases, complexity will also occur between the nucleic acid chains of different sources, forming a so-called hybridized double chain (heterod lexup), a process called hybridization.
can occur between DNA and DNA, between RNA and RNA, and between DNA and RNA. For example, a piece of natural DNA is interbred with a missing mutant of this DNA (assuming that several base pairs are lost in the middle of the DNA molecule), and small bubbles can be seen bulging in the middle of the hybrid double strand under an electron
scope
.
the location and length of the small bubble to determine where the missing mutation occurs and how much is missing. Nucleic acid hybridization technology is one of the common means to study nucleic acid structure and function, which can be used not only to test the absence and insertion of nucleic acids, but also to examine the common sequences and sequences of different biological species in nucleic acid molecules to determine their evolutionary relationship. Its application, of course, goes far beyond determining the location of mutations (Figure 15-10II.). I. Transsexuality, complexity and hybridization. The thick lines represent different DNA. A is the identification of hybrid and II. mutants. B stands for natural DNA; C is a missing mutant of B; a dotted wireframe is a missing part; D is a very useful technique for research and diagnosis developed on the basis of nucleic acid hybridization, showing that small bubbles are bulging out of natural DNA strands III. Thick lines represent probes, and Xs on the coarse line represent radioactive markers A single chain of a small segment (e.g. a dozen to hundreds) of nucleotide polymers with radioactive
isotopes
such as 32P, 35S, or biotin marking their ends or full chains can be used as probes. The DNA to be tested is denatured and adsorbed to a special membrane, such as a cellulose nitrate membrane.
the membrane and the probe together for a period of time, so that hybridization occurs. Rinse the film with a buffer. Because this membrane can more firmly adsorb double-stranded nucleic acids, single-stranded in flushing is washed away. A radioactive probe remains on the membrane if it binds to the DNA to be tested into a hybrid double strand. It is icing on whether the probe interbred with the DNA being measured by the radiological self-development of isotopes or the chemical color of biotin.
hybridization indicates that the DNA under test is homogeneity with the probe, i.e. the base sequences of the two can complement each other. For example, to know if a virus is related to a tumor, the virus's DNA can be made into a probe.
dna is extracted from tumor tissue and cross-breeded with the probe, the appearance of hybridized double strands indicates esoxuality between the two DNA. This does not mean that the virus causes tumors, but at least this is an important clue that can be further studied. probe technology has been applied in the diagnosis of genetic diseases. For example, the diagnosis of thalassemia or hemoglobin disease can extract DNA from the white blood cells of a confirmed patient, which is the diagnostic probe. With diagnostic probe examination, not only can the patient with symptoms be diagnosed, but also can find some asymptomatic recessive genetic diseases. A small amount of DNA can also be extracted from the fetal amniota.
because the probe technology is more sensitive, it makes the prenatal diagnosis of genetic diseases easier to do. Hybridization and probe technology are the basis of many
molecular biology
technologies and are increasingly used in biological and medical research as well as in clinical diagnosis.


. .