Direct electrophoretic detection of the allelic state of single DNA molecules in human sperm by using the polymerase chain reaction

Quantitative Polymerase Chain Reaction (Q-PCR) aims at determining the initial quantity of specific nucleic acids from the observation of the number of amplified DNA molecules. The most widely used technology to monitor the number of DNA molecules as they replicate is based on fluorescence chemistry. Considering this measurement technique, the observation of DNA amplification by PCR contains intrinsically two kinds of variability. On the one hand, the number of replicated DNA molecules is random, and on the other hand, the measurement of the fluorescence emitted by the DNA molecules is collected with some random error. Relying on a stochastic model of these two types of variability, we aim at providing estimators of the parameters arising in the proposed model, and, more specifically, of the initial amount of molecules. The theory of branching processes is classically used to model the evolution of the number of DNA molecules at each replication cycle. The model is a binary splitting Galton-Watson branching process. Its unknown parameters are the initial number of DNA molecules and the reaction efficiency of PCR, which is defined as the probability of replication of a DNA molecule. The number of DNA molecules is indirectly observed through noisy fluorescence measurements resulting in a so-called Hidden Markov Model. We aim at inference of the parameters of the underlying branching process, and the parameters of the noise from the fluorescence measurements in a Bayesian framework. Using simulations and experimental data, we investigate the performance of the Bayesian estimators obtained by Markov Chain Monte Carlo methods.

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Constructing Recombinant DNA Molecules by the Polymerase Chain Reaction

Current Protocols in Molecular Biology ◽

10.1002/0471142727.mb0317s24 ◽

1993 ◽

Vol 24 (1) ◽

Cited By ~ 1

Author(s):

Elaine A. Elion

Keyword(s):

Polymerase Chain Reaction ◽

Recombinant Dna ◽

Dna Molecules ◽

Chain Reaction ◽

Polymerase Chain

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Wx gene polymorphism in winter triticale collection samples

Agrobìologìâ ◽

10.33245/2310-9270-2020-157-1-80-87 ◽

2020 ◽

pp. 80-87

Author(s):

O. Levchenko

Keyword(s):

Polymerase Chain Reaction ◽

Gene Polymorphism ◽

Starch Content ◽

Infrared Spectrometry ◽

Wild Type ◽

Chain Reaction ◽

Winter Triticale ◽

Allelic State ◽

Wx Gene ◽

Polymerase Chain

The purpose of the study was to identify the collection of winter triticale in the allelic state of the waxi-genes and to identify sources with the presence of waxi-alleles for these genes. The surveys were conducted over 2017–2019 at the NSc Institute of Agriculture. The subject of the research are 43 collection samples of winter triticale, 29 of which are numbers of own breeding, 14 – breeding varieties of the National Institute of Agriculture of NAAS (9) and scientifi c institutions of Poland (1) and the Russian Federation (4). For control, we used soft winter waxy-wheat Sofi yka and wheat with wild of starch Oksana. Field, laboratory (infrared spectrometry, light microscopy, polymerase chain reaction (PCR)) methods, weights and mathematical and statistical methods of research were used to evaluate the collection material. According to the results of molecular genetic analysis of the Wx gene polymorphism in the winter triticale collection samples, it was found that all the tested samples had wild type alleles according to the Wx-B1 gene and were characterized by the absence of the Wx-D1 gene. The Wx-A1 gene revealed samples with both wild-type alleles and presence in the genome of the wax-allele. 8 collections with Wx-A1 gene alleles were selected: selection numbers 141, 153, 201, 223, 229 and varieties Lubomir, Petrol and Poliskii 7. The selected samples varied signifi cantly in terms of such characteristics as grain productivity, weight of 1000 grains, starch content. The tendency to decrease the size of the granules and increase the evenness of the granulometric structure of the starch in the samples with the presence of the wax-allele of the Wx-A1 gene was established. Wx-A1 gene allele samples are valuable starting material for the creation of new winter triticale varieties with increased amylopectin starch suitable for bioethanol processing. Key words: winter triticale, bioethanol, starch, polymerase chain reaction, amylopectin, amylose, allelic state of wax genes, waxi-allele, wild type.

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