China
Africa
Explore chapters and articles related to this topic
Molecular Approaches Towards the Isolation of Pediatric Cancer Predisposition Genes
Published in John T. Kemshead, Pediatric Tumors: Immunological and Molecular Markers, 2020
When total genomic DNA is “cut” with a particular enzyme, a heterogeneous population of characteristic restriction fragments is produced. Each fragment has a fixed length, the restriction fragment length (RFL). Agarose gels can be used to separate these fragments according to size. Using standard techniques, developed by Southern5 and Rigby et. al.,6 radioactive DNA probes can be used to hybridize with, i.e., “recognize”, homologous sequences within the heterogenous population. This homology is manifested as a band (or set of bands)on an autoradiograph at one or more specific locations determined by the size of the fragment being recognized (Figure 1). If there is no sequence variation between individuals in the region of the chromosome recognized by the probe, a consistent band pattern is produced at the same position on the gel. However, any variation in DNA sequence that affects a given restriction site, such that an existing one is removed or a new one is created, will generate variations in the band pattern observed. Since the bands produced are polymorphic, the variation is referred to as a restriction fragment length polymorphism (RFLP). This principle is illustrated in Figure 2. If an individual is heterozygous for the polymorphic variants, then both of the individual homologous chromosomes can be identified (see Figure 2). Differences in DNA sequence at the same locus are allelic and the bands produced on the autoradiograph are conveniently referred to as alleles.
The Genetics of Alzheimer Disease:
Published in Robert E. Becker, Ezio Giacobini, Alzheimer Disease, 2020
Molecular genetic techniques allow creation of innumerable DNA markers by cleaving DNA using restriction endonucleases. The resulting DNA fragments, also known as restriction fragment length polymorphisms (RFLPs), can then be used as markers in linkage analyses. Since RFLPs of different lengths from any chromosome can be produced, microdissection of the human genome is possible. More extensive discussion of such techniques are reviewed in another chapter of this book (Wizniewski, 1989).
Immunology of Insulin-Dependent Diabetes Mellitus
Published in Irun R. Cohen, Perspectives on Autoimmunity, 2020
Christian Boitard, H. O. McDevitt
Characterization of the class II allelic gene products associated with IDDM have relied in the past on serological techniques. The associations defined are partial, and the majority of subjects with susceptibility alleles never develop IDDM. In addition, 10% of IDDM patients do not carry the susceptibility alleles defined by serological markers. Diseases which do not appear to be related to IDDM are associated with identical susceptibility alleles. Serological definition of susceptibility alleles, therefore, may be incomplete and/or insufficient. Restriction fragment length polymorphism (RFLP) is a first step in characterizing, at the DNA level, susceptibility alleles and in verifying whether alleles which have different amino acid sequences but share identical antigenic determinants are detected as different by RFLP. This approach has been developed using a DR β probe to study HLA-DR in DR4 and DR3 heterozygous individuals. An increased frequency of a Pstl 18-kilobase (kb) fragment has been observed.176 Similar studies in DR2 healthy and diabetic individuals using a DC β cDNA probe have also allowed the definition of a 2.2-kb fragment which may be related to protection against IDDM.177
Cytogenetic and molecular genetic methods for chromosomal translocations detection with reference to the KMT2A/MLL gene
Published in Critical Reviews in Clinical Laboratory Sciences, 2021
Nikolai Lomov, Elena Zerkalenkova, Svetlana Lebedeva, Vladimir Viushkov, Mikhail A. Rubtsov
The Southern blot [43,44] was the first method to detect changes in genomic DNA sequences detected by restriction fragment length polymorphisms (RFLPs). In this method, total genomic DNA is cleaved with restriction endonucleases. The obtained fragments are then separated in an agarose gel, transferred to a nylon membrane, hybridized with labeled probes, and finally, the bound probes are detected. This method has been used to determine the rearranged genes in many AL-associated recurrent chromosomal translocations (see the analysis of the chimeric KMT2A-AFF1 gene in t(4;11)(q21;q23) [45]). Southern blot has also been utilized for the detection of rearrangements in diagnostic samples as the length of restricted DNA fragments differ in rearranged and wild-type alleles [46].
The relationship between KRAS LCS6 polymorphism and endometrium cancer
Published in Journal of Obstetrics and Gynaecology, 2020
Feyza Nur İncesu Çintesun, Özlem Seçilmiş Kerimoğlu, Ersin Çintesun, Süleyman Nergiz, Hasan Acar, Çetin Çelik
The Restriction Fragment Length Polymorphism (RFLP) method is a method which allows the examination of DNA profiles formed by separation in the agarose gel electrophoresis system of DNA segments and fragmentation with a restriction endonuclease enzyme specific to genomic DNA. By identifying specific sequences in the DNA of these enzymes, which are found in bacteria, the fragmentation process occurs in the identified region or in another specific sequence outside this region. Genotyping of single nucleotide polymorphisms occurs in two steps with this method. The first step is based on PCR product sequencing or enzyme fragmentation after gel electrophoresis for the genotyping of the polymorphic region by hundreds of thousands of proliferations with PCR of approximately 200 base pairs around the single nucleotide polymorphism (SNP) in the DNA region (Linn and Arber 1968).
Diagnostic markers for glaucoma: a patent and literature review (2013-2019)
Published in Expert Opinion on Therapeutic Patents, 2019
Mutations in the myocilin gene called p. Q368X were detected by restriction fragment length polymorphism (RFLP) analysis. RFLP exploits variations in homologous DNA sequences, known as polymorphisms, to distinguish individuals, populations, or species, or to pinpoint the locations of genes within a sequence. In RFLP analysis, a DNA sample is digested into fragments by one or more restriction enzymes, and the resulting restriction fragments are then separated by gel electrophoresis according to their size. Undoubtedly, RFLP analysis has been an important early tool for genome mapping, localization of genes for genetic disorders, determination of risk for disease, and paternity testing. However, restriction analysis is a relatively expensive and time-consuming method, which might last over 3 days. As a result, faster and cheaper DNA sequencing technologies have been developed nowadays which made RFLP obsolete.