Random Amplified Polymorphic DNA (RAPD) is J. Williams and J. Welsh's two research groups also proposed a random primer amplification in 1990 to search for genetic markers for polymorphic DNA fragments. It is based on PCR technology and uses random oligodeoxynucleotides as primers for PCR reactions. Amplification of gene DNA reveals DNA mapping and genetic relationship with species, identification of phylogenetic molecular levels, and molecular biology and molecular ecology. An obvious feature of the RAPD marker is that RAPD primers are non-specific. A set of discrete DNA fragments can be obtained by PCR amplification using genomic DNA of unknown sequence as a template. The RAPD primers required are short, with about 10 oligos. Nucleotide can be. Second, a set of primers can be used for different organisms, and a set of standard primers can be established for the identification of polymorphisms in biological species. Due to the use of the DNA amplification instrument, the automation of the operation is high and the amount of analysis is large, and steps such as preparing a probe, isotope labeling, Southern blotting, and molecular hybridization in the RFLP are eliminated, thereby forming a rapid RAPD analysis and the required DNA. Advantages such as less samples: Especially the PAPD markers can be used to construct genomic DNA fingerprints of species without any research on molecular biology. I. Application of RAPD technology in insect taxonomy Traditional methods of insect taxonomy are based on external morphology. Within a large taxonomic unit, the taxonomic status of a species can be clearly defined. However, for some species, it is determined. When the taxonomic status of a genus, clan, or species is often determined by its external form, it is often difficult to determine. Since the evolution of species is essentially the evolution of the genome, molecular taxonomy can more accurately and even quantitatively analyze the evolutionary speed, the genetic distance, and the inference of system relationships at the DNA level. In recent years, the DNA taxonomy has made great progress and has received more and more attention from people. It has become an important supplement to classical classification methods. Therefore, by selecting suitable primers, RAPD technology can easily display the differences within the genes, thus providing a basis for accurately determining the taxonomic status of species. Black et al. (1992) first applied RAPD technology to the identification and comparison of four aphids. They used four 10-base random primers to conduct RAPD reaction on four aphids to detect the amplification product polymorphism. The results showed that According to the electrophoretic bands, 4 species can be distinguished. At the same time, the polymorphisms of the amplified products in different species within the species and in different individuals in the same biotype were detected, and the polymorphisms of the amplification products among different individuals within the population. Puterka et al. (1993) systematically analyzed Diuraphic noxia Mordvilko, a Russian wheat moth, collected in several countries using RAPD technology. Seven primers yielded 69 polymorphic bands. This method can identify all the tested populations. At the same time, the cluster analysis showed that the populations collected from South Africa, Mexico, France, and Turkey were similar to the U.S. population, and the genetic relationship was very close, while the populations in the Middle East and southern Russia had large variation and distant genetic relationships. Yang Xiaowen et al. (1999) used RAPD technology to study the DNA polymorphisms of haze on peaches, canola and tobacco. The results showed that, compared with yellow-green, the color of haze was closer to red and brown. Han Yali (2001) used a random amplified polymorphic DNA technique to compare the polymorphisms of the genomic DNA of 3 species and 6 species of Aphid in the family Polyptera, and constructed a UPG clustering diagram based on the degree of fragmentation. The results show that the sharing degree of the fragments of the big-legged blubber and the cane-winged dragonfly is 0.375. The fragment sharing degree of the red-winged knees and the drum-winged knees was 0.268, and the fragment sharing degree between the Asian Carriage and the Carassius auratus was also 0.286. Second, the application of RAPD technology in insect molecular ecology research Insect molecular ecology is to study the relationship between biology and biology and environment from the genetic level. At present, RAPD technology is mainly used in this field for genetic variation of populations and genetic differentiation among populations, identification and identification of natural enemy insects, identification of pest biotypes, individual genetic markers to identify genetic relationships, and research on insect reproductive strategies. Roehrdanz et al. (1993) used RAPD technology to identify the natural enemies of the North American ladybird beetle of different geographical origins, which played a decisive role in the correct introduction of natural enemies and the control of aphids, and pointed out that the RAPD technology can be used to distinguish the natural enemies that are close relatives. There is great economic significance in using natural enemies to control pests. Chen Naizhong (1997) identified four species of warehouse larvae using RAPD technology. The results of electrophoresis showed that five primers produced amplified bands that could be used to distinguish these four insects, indicating that RAPD technology is particularly useful for insects, especially larvae. There are application prospects in rapid identification. Gawel et al. (1993) extracted DNA from Aleyrodinae eggs and nymphs for RAPD amplification. The 20 primers used can accurately distinguish the two biotypes (A and B) of Amygdalus. . Ban-o et al. (1997) reported that biotype B of Bemisia inconspicua (Quaintance) can be distinguished from other biotypes using RAPD technology, indicating that RAPD technology can be used to distinguish closely related biological groups. Wilkerson (1993) used RAPD markers to study two experimental populations of Anopheles mosquitoes in Africa. Using 57 primers, 377 DNA bands were amplified. There were 295 bands between the two populations; 13 primers with clear bands and good repeatability were selected to further amplify the DNA of 30 individuals of each species, among which 7 bands were amplified with different bands. , indicating that RAPD technology can effectively detect the variation that occurs within the population. Blanchetot (1992) used M13 bacteriophage DNA probes and globin gene repeat probes to study reproductive strategies of Megalle rotundata, demonstrating that the offspring in each nest basically come from a single female, and in most cases Females only mate with a single male. Lu (1994) applied the technique of DNA repeat sequence tagging to the study of Spodoptera fragiperda. There are partial overlaps in the distribution areas of the two populations of the species, one of which is infested with corn and the other is based on rice and feed forage. The results show that the DNA repeat sequence labeling technique can completely separate two morphologically indistinguishable populations. At the same time, the migration, population structure and reproductive behavior of the autumn armyworm population were also studied. Zhao Zhongming et al. (2001) used RAPD technology to analyze the genetic variation of 12 geographic populations of five sister species of Drosophila melanogaster. There were 30 primers in 40 kinds of 10 bases random primers and satisfactory amplification results were obtained for each population. Among 161 RAPD markers, 129 were polymorphic, and this composite species was constructed by UPGMA method. The clustering diagram. 3. The application of RAPD technology in pest resistance has been in the past 20 years. In order to control the damage of pests, a large number of pesticides are applied to crops. Many pests have developed a strong resistance to drugs. The emergence of drug resistance is the result of genetic mutations and selections that are associated with resistance to genetic mutations that occur in the population under conditions of constant external loop resistance. Due to the imperfect theory of pest genetics and the lack of diagnostic techniques for resistance mutants, previous researches on resistance mechanisms were mostly limited to physiology, and they rarely reached the molecular level. Once RAPD technology emerged, it was rapidly used in the research of the resistance mechanism of pests to detect the genetic variation of resistance. Even if people lack a deep understanding of the related resistance genes, they can rely on the DNA linked by three resistance genes. Markers identify the genetic variation of resistance in the population. Ruan Changhui et al. (1996) used RAPD technology to analyze the inheritance pattern of cyhalothrin resistance in Helicoverpa armigera. A total of 47 DNA bands were amplified from the R and S parents by screening three random primers. Up to 27 bands were selected; three RAPD molecular markers related to anti-cyhalothrin were initially screened, namely OKG4-1300, OPG6-1450, and OPG8-535, which can occur simultaneously in the R parent and the F1 generation in the forward and reverse crosses. It does not appear in the S parent, and is consistent with the results of the genetic method of resistance, demonstrating the reliability of this method. Raymond (1991) used this technique to study the mechanism of the development and spread of resistance to effective phosphorus pesticides by Culex pipiens (Culexpipiens), demonstrating that the spread of the esterase B2 gene that causes the resistance of Culex mosquitoes has a single origin and spreads through migration. Different regions. IV. Application of RAPD technology in crop pest resistance RAPD and RAPD markers are two different concepts. The DNA fragments obtained by RAPD amplification have been identified to be useful as molecular markers, and they are called RAPD markers. Myburg et al. (1998) carried out the linkage marker study of Dn2 gene of wheat against Russian wheat aphid, and detected a total of 2 700 loci. Among them, 4 RAPD fragments were identified as markers linked to the Dn2 gene. The isolation of the F2 resistance gene indicated that the four fragments were closely linked to the Dn2 gene, and these four fragments were used as probes for Southern analysis to confirm their homology. Blanchetot (1993) used RAPD and RFLP techniques to study the genetic dependencies between African vulgaris and intermediate host African tsetse fly. And the African tsetse inbred lines and gene linkage maps provide the basis for the design of trefoil flies biological control targets in Africa: Roche et al. identified the resistance through the molecular marker study of apples resistant to Dysaphis devecta. Three RFLP markers and four RAPD markers linked by sex genes (sdl). And they were located. The acquisition and localization of these markers have good prospects for use in resistance breeding. Heckel et al. used 117 random primers to carry out RAPD marker research on the genomic DNA of Plutella xylostella anti-Bt strains and susceptible strains. Of the 223 amplified bands obtained, 105 were unique to susceptible strains and 118 were unique to resistant strains. In addition, Liu Chunyu et al. performed RAPD assays on the silkworm and ramie silkworm hybrid progenies. Although this crossbreeding did not occur in male and female nuclear fusion, it caused a change in the maternal genome structure in the amplified product of the hybrid offspring. Not only there are a large number of amplification bands that are identical to the mother, but also a number of "variant bands" that are different from the mother. The emergence of this variant band may be due to the fact that the ramie silkworm sperm carries foreign DNA into the eggs of the silkworm, resulting in rearrangement of the original genetic structure of the maternal genome (including deletions, substitutions, insertions, etc.). This - the study has carried out a beneficial exploration in the detection and tracking of insect foreign genes. V. Analysis of Insect Genomes and Construction of Gene Maps With the continuous development of molecular biology techniques, especially the discovery of RAPD technology with low cost and simple operation, it has become possible to construct insect gene maps at the molecular level. Hunt et al. (1995) constructed a genetic map of honeybees using RAPD markers. The map includes 26 linkage groups with a total length of 161,720 kb (estimated beeline genome is 24,150 kb in length) and an average of 10 polymorphic loci screened by 10 bp primers. Up to 2.8, and based on this, the sex-determining X locus, the genes controlling the black body color, and the malate dehydrogenase gene were respectively located on different linkage groups. He Ningjia et al. (2001) constructed a linkage map of the silkworm with the combination of SADF and RAPD markers. 40 selective amplification of polymorphic primers and 137 random amplified polymorphic primers were used to screen the Bombyx mori F2 population. . The application of RAPD technology in entomological research has only just begun. With the deepening of insect molecular biology research and the development of RAPD and related technologies, RAPD technology will be combined with other technologies to find, modify and utilize insect genes. New countermeasures for pest management and new ways to protect and improve beneficial insects will play an important role. Chen Qing, World Agriculture
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