China
Africa
Explore chapters and articles related to this topic
Genetics of Endocrine Disorders and Diabetes Mellitus
Published in George H. Gass, Harold M. Kaplan, Handbook of Endocrinology, 2020
Bess Adkins Marshall, Abby Solomon Hollander
The most accurate way to establish prenatal diagnosis of 21-hydroxylase deficiency is to do molecular genetic studies on amniotic or chorionic villus samples. Extracted DNA can be evaluated by Southern blot analysis.20,21 Mutations in the 21-hydroxylase gene can be detected using polynucleotide probes designed to detect mutations in CYP21B. Through the use of PCR, amplified DNA can be obtained from very small samples.22 Although the reports of direct molecular genetic testing to establish prenatal diagnosis of 21-hydroxylase deficiency contain small numbers of patients, the results are uniformly accurate.20–22
Practical Approach to Molecular Biology in Hematopathology
Published in Harold R. Schumacher, William A. Rock, Sanford A. Stass, Handbook of Hematologic Pathology, 2019
Anwar Mikhael, Harold R. Schumacher
In theory, polymerase chain reaction (PCR) allows the amplification of DNA from a single cell. This technique can reproducibly detects cells present at a concentration of 1 to 104 to 105. Because of this sensitivity, PCR has in many instances replaced the use of Southern blot in diagnostic hematopathology (Table 5). More important, it can be applied to small tissue samples (skin biopsies) and to samples with substantial DNA degradation, which are often not suitable for Southern blot technology.
A Survey of Newer Gene Probing Techniques
Published in Victor A. Bernstam, Pocket Guide to GENE LEVEL DIAGNOSTICS in Clinical Practice, 2019
Southern blot analysis in its various modifications may still be used for clinical molecular genetic diagnosis for some time to come, despite the revolutionary developments in the amplification of diagnostic targets.
An insight into clinical and laboratory detections for screening and diagnosis of cervical cancer
Published in Expert Review of Molecular Diagnostics, 2023
Shruthi Padavu, Pooja Aichpure, Ballamoole Krishna Kumar, Anoop Kumar, RadhaKanta Ratho, Shipra Sonkusare, Indrani Karunasagar, Iddya Karunasagar, Praveen Rai
Southern blotting involves electrophoresing HPV DNA before transferring it to a membrane film and hybridizing it into a tagged probe. Southern blot analysis has several advantages: it allows the confirmation of the HPV type identified; it reduces the risk of false negatives by detecting quantities of DNA that are often undetectable when stained with ethidium bromide (EtBr); and, finally, it lowers the possibility of false positives by differentiating minor variations in size between two PCR products [54]. However, Southern blot analysis is labor-intensive and requires significant amounts of DNA that may not be achievable from small tissue biopsies. The test’s drawbacks include its inability to detect episomes with low copy numbers [55]. This method is time-consuming, expensive, and requires expertise [56,57]. Although the Southern blot was once thought to be the standard method for detecting HPV DNA, it has not been used for routine diagnosis in healthcare settings [53].
T cell receptor revision and immune repertoire changes in autoimmune diseases
Published in International Reviews of Immunology, 2022
Ronghua Song, Xi Jia, Jing Zhao, Peng Du, Jin-an Zhang
The limited understanding of autoantigens and autoreactive TCRs once stopped people from further exploration of AIDs. Analyzing TCR repertoire is a very useful approach, but is also challenging due to the large number and highly variable nature of TCR repertoire. Despite deciphering the entire T-cell repertoire in diseases is still far away, important progresses have been made to bring us closer to this goal. Scientists have tried a variety of methods to detect the TCR library, such as Sanger sequencing, flow cytometry, PCR for CDR3 region, and Southern blot for TCR DNA [47–50], but all these methods have their own limitations. Sanger sequencing can only sequence up to 700 bp nucleic acid, and its low-throughput makes it a laborious method with high failure rate [49]. Flow cytometry has the advantage of assessing actual protein expression on cell surface, but it can only reveal the frequency of V-family usage [48]. Multiplex PCR, an RNA (cDNA)-based approach was also used in TCR detection [50]. However, this method can hardly make precise and credible comparisons between samples, even when using a complex scoring system that defines the heights of clonal peaks relative to the remaining repertoire [50,51]. Additionally, the unavoidable PCR bias, caused by differences in primer annealing and amplification conditions, also hindered its application [51]. Southern blot is currently not used anymore, because it requires large amounts of DNA and is not effective or accurate [47].
Telomeres as a molecular marker of male infertility
Published in Human Fertility, 2019
Ewa Boniewska-Bernacka, Anna Pańczyszyn, Natalia Cybulska
In the Southern blot method genomic DNA is subjected to restriction endonuclease digestion, most often HinfI/RsaI, at sites that do not contain telomere sequences. The digested DNA is electrophoretically separated in agarose gel (Aviv et al., 2011), transferred to a nylon membrane and then hybridized to specific probes (Law & Lau, 2001). This method is used to determine the average length of telomeres; however, it is not suitable for analysis of short telomere sequences. The Southern blot method does not require specialized equipment; however, it requires time and experience to analyze the results. The disadvantages of this method include possibility of erroneous determination, which may be due to the occurrence of sequence-changing polymorphisms at restriction enzyme sites. This method is considered to be a reference method for validating newly introduced techniques (Kimura et al., 2010; Zong, Wang, Chen, & Cui, 2014).