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1
artículo
The avocado (Persea americana Mill) is a plant food interest, the problem at the level of the crop are diseases that reduce both plant and post-harvest quality; among them are produced by the viroids, including Avocado you have to sunblotch viroid (ASBVd) and Potato spindle tuber viroid (PSTVd) who dramatically reduce the organoleptic properties of the fruit. During the production process of said culture is needed pathogen free seedlings necessitating early detection. Currently there are methods of detecting an estimated cost of 24 USD through the use of real-time PCR technique.In the search for methods cheaper and reliable detection it was carried out standardization of a conventional PCR protocol that allows reducing the cost of detecting viroid in avocado mother plants. To this was done using the technique of reverse transcriptase reverse coupled the polymerase chain reaction (RT-PCR)...
2
tesis de grado
Para facilitar la identificación rápida y especifica de Campylobacter jejuni se desarrolló un Test de Reacción en Cadena de la Polimerasa (PCR) a partir de muestras de heces de niños, con problemas gastrointestinales, menores de 24 meses de edad. Los cebadores (primers) elegidos, CAMPYJ y CAMPYJ2, para la realización del PCR se diseñaron sobre la base de una secuencia específica del gen MapA, este gen codifica a la proteína A (24KD), que es parte estructural de la membrana celular, presente en todas las cepas de Campylobacter jejuni. Para la extracción de ADN directamente de heces se evaluaron 04 protocolos diferentes, mostrando el Método de Franket y col. un ADN genómico más puro y conservado. Para la estandarización del PCR se utilizaron cepas de Campylobacter jejuni, cuyo límite de detección con el par de cebadores fue 15 bacterias, equivalente a 100 fentogramos de ADN...
3
artículo
The present paper describes a protocol applied to fecal samples of cattle for the isolation and purification of Cryptosporidium sp. oocysts and the possible use of the same ones for genetic studies by means of PCR.
4
artículo
The present paper describes a protocol applied to fecal samples of cattle for the isolation and purification of Cryptosporidium sp. oocysts and the possible use of the same ones for genetic studies by means of PCR.
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artículo
The aim of the study was to identify serotypes of suspicious isolates of Salmonella spp from breeding guinea pigs in samples collected within the first week of parturition to detect animals carrying the bacteria. The guinea pigs were clinically normal and reared in a commercial farm in Pachacamac, Lima, Peru. The farm was free of Salmonella outbreaks in the last four years. A total of 272 paired samples consisting in rectal and vaginal swabs per animal were collected and analyzed using standardized microbiological protocols. The DNA was extracted from suspected isolates of Salmonella sp and then these samples were analyzed by multiplex PCR to detect the presence of invA, prot6E and fliC genes which are specific for Salmonella spp, Salmonella Enteritidis and Salmonella Typhimurium respectively. Eight animals (12 swabs) were positive to Salmonella spp. All 12 isolates amplified invA (Salmo...
6
artículo
The aim of the study was to identify serotypes of suspicious isolates of Salmonella spp from breeding guinea pigs in samples collected within the first week of parturition to detect animals carrying the bacteria. The guinea pigs were clinically normal and reared in a commercial farm in Pachacamac, Lima, Peru. The farm was free of Salmonella outbreaks in the last four years. A total of 272 paired samples consisting in rectal and vaginal swabs per animal were collected and analyzed using standardized microbiological protocols. The DNA was extracted from suspected isolates of Salmonella sp and then these samples were analyzed by multiplex PCR to detect the presence of invA, prot6E and fliC genes which are specific for Salmonella spp, Salmonella Enteritidis and Salmonella Typhimurium respectively. Eight animals (12 swabs) were positive to Salmonella spp. All 12 isolates amplified invA (Salmo...
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artículo
The purpose of this study was to select for the extraction of DNA from Leptospira spp from urine samples for the diagnosis of bovine leptospirosis by PCR. Three methods of DNA extraction methods were used: Ethanol-Sodium Hydroxide (EtNa), Chelex® 100 chelating resin and the PureLink® Genomic DNA Mini Kit commercial extraction case. The extracted DNA served for the standardization of three PCR protocols for the identification of the rrl, hap1 and rrs genes, respectively, in the AGROCALIDAD, Ecuador laboratory. A total of 72 bovine urine samples were collected from livestock farms in the province of Manabí, Ecuador. Ten positive samples were obtained by amplifying the rrl gene, which identifies the genus Leptospira. Of the genus-positive samples, eight amplified for the hap1 gene, which codes for the main outer membrane protein of pathogenic species. The agreement between the DNA extrac...
8
artículo
The purpose of this study was to select for the extraction of DNA from Leptospira spp from urine samples for the diagnosis of bovine leptospirosis by PCR. Three methods of DNA extraction methods were used: Ethanol-Sodium Hydroxide (EtNa), Chelex® 100 chelating resin and the PureLink® Genomic DNA Mini Kit commercial extraction case. The extracted DNA served for the standardization of three PCR protocols for the identification of the rrl, hap1 and rrs genes, respectively, in the AGROCALIDAD, Ecuador laboratory. A total of 72 bovine urine samples were collected from livestock farms in the province of Manabí, Ecuador. Ten positive samples were obtained by amplifying the rrl gene, which identifies the genus Leptospira. Of the genus-positive samples, eight amplified for the hap1 gene, which codes for the main outer membrane protein of pathogenic species. The agreement between the DNA extrac...
9
artículo
In 2019, in the city of Wuhan, China, patients were diagnosed with a novel coronavirus originally named 2019-nCoV, currently known as SARS-CoV-2. The alarming expansion of this pandemic makes it necessary to find out and implement reliable diagnostic methods in order to properly detect and treat patients, thus contributing to slowing down the spread of the disease. It is extremely important to have trained professionals in sample collection, good clinical laboratory practices, and molecular and hematological techniques to adequately detect any cases of infection with this virus. Moreover, standardized protocols are essential for obtaining, transferring and storing samples to stop the spread of the COVID-19 pandemic caused by SARS-CoV-2.
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artículo
Hookworm is generally caused by Necator americanus (Stiles, 1902), causing digestive symptoms and anemia. The diagnosis by coprology can have low sensitivity, especially in light parasite loads. The Polymerase Chain Reaction (PCR) is a sensitive and specific technique, which must be adapted to laboratory conditions and positive controls are necessary. The objective of this work was the standardization of the PCR technique for the amplification of the ITS-1 sequence of N. americanus in stool samples and its cloning for its use as a control. Three DNA extraction protocols were standardized (Phenol/chloroform, Saline Precipitation, and Chelex® 100 Resin). Optimal reagent concentrations (MgCl , BSA, dNTP, primers, and Taq polymerase) were determined, as well as the hybridization 2 temperature and number of cycles. The analytical sensitivity and specificity of the technique was determined. F...
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artículo
Nuestro país es uno de los principales exportadores de chirimoya en el mundo, siendo Lima la región con mayor producción y exportación. Actualmente, existe poca información genética acerca de la identificación de la especie Annona cherimola Mill “chirimoya” mediante el uso de código de barras de ADN, herramienta molecular que permite tener un registro correcto en la identificación precisa de la especie. El objetivo de este estudio fue optimizar la técnica de extracción y amplificación por PCR como parte de la identificación molecular de Annona cherimola Mill “chirimoya”, mediante el uso de dos marcadores universales como matK y rbcL relacionados al gen de maduración y fotosíntesis. Se analizaron un total de 154 hojas colectadas de chirimoya extraídas de 26 árboles, distribuidas en 6 zonas de la región Lima-Perú (Ate, Callahuanca, Carabayllo, La Molina, Lima y T...
12
artículo
A systematic review of the diagnostic methods described in the literature for the identification and characterization of species of the genus Salmonella was carried out. The review was carried out using publications from 2015 to 2021 registered in the NCBI/PubMed, ScienceDirect, Scopus, SciELO, Google Scholar and Springer databases and applying the PRISMA methodology. In total, 61 articles were analysed, grouping the data according to country of origin, type of sample, technique, microbiology protocol and genes used in molecular tests. The microbiology protocols are based on performing pre-enrichment in peptone water and enrichment in tetrathionate (TT) and Rappaport-Vassiliadis (RV) broths, especially with faecal or environmental samples, to subsequently carry out isolations in selective and differential media. The identification of some Salmonella sp serovars that cause disease in poul...
13
tesis de grado
Con la finalidad de insertar los genes cry3Ca1 y cry7Aa1 en Ipomoea batatas (L.) Lam. cv. “Huachano” para conferir resistencia a Cylas puncticollis y C. brunneus “gorgojos del camote”, se desarrolló la transformación genética mediada por Agrobacterium tumefaciens, siguiendo dos protocolos de regeneración y transformación. Partiendo de 46 meristemos y 282 hojas con peciolo, se obtuvieron 8 eventos transgénicos de inserción completa del ADN - T (4 por cada protocolo de transformación) y un evento de inserción incompleta. Con una eficiencia de transformación de 8.70%, el protocolo a partir de meristemos demostró ser más eficaz en la obtención de eventos transgénicos de inserciones completas que el protocolo a partir de hojas con peciolo, con el cual se obtuvo una eficiencia de transformación de 1.42%. A su vez, los regenerantes fueron evaluados mediante pruebas in vit...
14
tesis de grado
El dengue es una enfermedad producida por el virus del dengue. En la actualidad existen diversas técnicas que son utilizadas para su diagnóstico, sin embargo, la RT-qPCR permite obtener resultados confiables y rápidos con una alta sensibilidad y especificidad. Además, a diferencia de la prueba ELISA NS1, nos permite identificar los serotipos del DENV. El objetivo del presente Trabajo de Suficiencia Profesional fue diseñar una propuesta para la implementación de la técnica RT-qPCR para la detección molecular temprana del DENV y sus serotipos en el Laboratorio Referencial de Salud Pública – DIRESA JUNÍN. Se revisó la bibliografía disponible sobre los primers y las sondas que están siendo utilizados para el diagnóstico del DENV y sus serotipos. Estas secuencias fueron evaluadas mediante la herramienta “Nucleotide-BLAST” y en base a los resultados obtenidos se propusieron...
15
artículo
Molecular research demand the creation of new protocols and permit the obtain a high quality DNA in the most quantity of species and know the genetic diversity, for that objective of the present research was obtained the purification and amplification of scales DNA in the snake Bothrops atrox. the scales were recollected in the field using the visual encounter survey [5] The For the purification of the DNA was modification the protocol [4], the DNA purity was verifiable for the electrophoretic method and it quality and concentration for spectrophotometry, the amplification was carried for Polymerase chain reaction-PCR using random primers. The results show the successful DNA purification, showing an acceptable quality ratio (2) and concentration (585.85 µg/mL), at the same time was made the amplification of DNA for PCR. We conclude indicating the useful of this technique for the purific...
16
tesis de grado
La difilobotriosis es una zoonosis causada por la ingestión accidental de la larva plerocercoide de Diphyllobothrium, familia Diphyllobothridae, que se encuentra enquistada o libre en la superficie visceral de los peces teleósteos marinos, considerados hospederos paraténicos o intermediarios. El presente trabajo tuvo como objetivo identificar molecularmente los estadios de plerocercoide y adulto de Diphyllobothrium sp. obtenidos de humanos, lobos marinos y peces en el Perú. Se muestrearon 3668 ejemplares de Engraulis ringens “anchoveta”, 22 de Sciaena deliciosa “lorna”, 6 de Sciaena callaensis “lorna grande”, 6 de Trachurus symmetricus murphyi “jurel” frescas y congeladas en diferentes periodos estacionales, adquiridos de diferentes Puertos del Norte (Chiclayo), Centro (Lima) y Sur (Ica) de la costa peruana. Los especímenes adultos de Otaria byronia se obtuvieron de ...
17
tesis doctoral
Universidad Nacional Agraria La Molina. Escuela de Posgrado. Doctorado en Agricultura Sustentable
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tesis de grado
The study of malaria in birds in these last two decades has been increasing, for this, rapid and low-cost protocols are being developed for the extraction of genomic DNA, amplification and screening, allowing the presence of these hemoparasites to be detected. The objective of this research was to compare genomic DNA isolation protocols and to identify birds parasitized by the Plasmodium, Haemoproteus and Leucocytozoon genera that cause avian malaria in the Loreto Region, for which the salt precipitation protocol was used and modified ( Salting-out) and the method of isolation of blood cells by density gradient according to the conditions of the collected sample, proceeding to amplify by means of the Polymerase Chain Reaction in real time (qPCR) obtaining as results that the salt precipitation protocol showed better results in terms of its concentration and quality of ratios compared to ...
19
tesis de maestría
Aeromonas salmonicida y Yersinia ruckeri son bacterias patógenas de Trucha arcoíris; su detección temprana mediante el uso de pruebas de diagnóstico es fundamental para evitar la propagación bacteriana y la mortalidad excesiva en los sistemas de producción. Sin embargo, el uso de pruebas de diagnóstico no se realiza debido a que son costosas, requieren de laboratorios sofisticados y se realizan en ciudades alejadas de los centros de producción. Por ello, este trabajo tuvo como objetivo estandarizar la técnica de Amplificación Isotérmica Mediada por Bucle (LAMP), una prueba de diagnóstico molecular rápido altamente sensible y de bajo costo, para detectar simultáneamente A. salmonicida y Y. ruckeri en cepas aisladas de Trucha arcoíris. Se diseñó primers LAMP para la detección de A. salmonicida y Y. ruckeri mediante los genes vapA y rucC-rupB, respectivamente. Se elaboró ...
20
artículo
La Ley Forestal y de Fauna Silvestre en el Perú, permite la caza de subsistencia de la fauna silvestre para autoconsumo y el comercio de pieles por las comunidades locales. Estas pieles son secadas en la modalidad de seco dulce y luego almacenadas en las viviendas de los pobladores hasta su comercialización. Por lo tanto, son altamente manipuladas y expuestas a efectos ambientales diversos. En el estudio se recolectó 300 muestras de pieles, procedentes de tres cuencas de ríos (Amazonas, Napo y Pacaya- Samiria) y se estandarizó un protocolo de extracción del ADN para pieles de Pecari, evaluándose dos métodos de extracción de ADN (Fenol– Cloroformo – Alcohol isoamilico y por columna). Se logró cuantificar el ADN con valores entre 164 y 3461 ng/µL obtenido por el Fenol– Cloroformo – Alcohol isoamilico. Asimismo, se estandarizó un protocolo de PCR para evaluar la eficienc...