title: Stem-Loop RT-qPCR as an Efficient Tool for the Detection and Quantification of Small RNAs in Giardia lamblia.
abstract: Stem-loop quantitative reverse transcription PCR (RT-qPCR) is a molecular technique used for identification and quantification of individual small RNAs in cells. In this work, we used a Universal ProbeLibrary (UPL)-based design to detect-in a rapid, sensitive, specific, and reproducible way-the small nucleolar RNA (snoRNA) GlsR17 and its derived miRNA (miR2) ofGiardia lambliausing a stem-loop RT-qPCR approach. Both small RNAs could be isolated from both total RNA and small RNA samples. Identification of the two small RNAs was carried out by sequencing the PCR-amplified small RNA products upon ligation into the pJET1.2/blunt vector. GlsR17 is constitutively expressed during the 72 h cultures of trophozoites, while the mature miR2 is present in 2-fold higher abundance during the first 48 h than at 72 h. Because it has been suggested that miRNAs inG.lambliahave an important role in the regulation of gene expression, the use of the stem-loop RT-qPCR method could be valuable for the study of miRNAs ofG.lamblia. This methodology will be a powerful tool for studying gene regulation inG. lamblia, and will help to better understand the features and functions of these regulatory molecules and how they work within the RNA interference (RNAi) pathway inG. lamblia.
authors.affiliation: CONACYT-Instituto Nacional de Pediatría, Secretaría de Salud, Ciudad de México 04530, Mexico. jmarcialqu@conacyt.mx.
authors.forename: Jaime
authors.initials: J
authors.lastname: Marcial-Quino
authors.affiliation: Laboratorio de Bioquímica Genética, Instituto Nacional de Pediatría, Secretaría de Salud, Ciudad de México 04530, Mexico. saulmanzo@ciencias.unam.mx.
authors.forename: Saúl
authors.initials: S
authors.lastname: Gómez-Manzo
authors.affiliation: Laboratorio de Ingeniería Genética y Metabolitos Secundarios, Departamento de Biotecnología, Universidad Autónoma Metropolitana-Iztapalapa, Ciudad de México 09340, Mexico. degfff@yahoo.com.
authors.forename: Francisco
authors.initials: F
authors.lastname: Fierro
authors.affiliation: Laboratorio de Neurociencias, Instituto Nacional de Pediatría, Secretaría de Salud, Ciudad de México 04530, Mexico. america_vc@yahoo.com.mx.
authors.forename: America
authors.initials: A
authors.lastname: Vanoye-Carlo
authors.affiliation: Laboratorio de Parasitología Experimental, Instituto Nacional de Pediatría, Secretaría de Salud, Ciudad de México 04530, Mexico. yadirg@gmail.com.
authors.forename: Yadira
authors.initials: Y
authors.lastname: Rufino-González
authors.affiliation: Colegio de Ciencias y Humanidades, Plantel Casa Libertad, Universidad Autónoma de la Ciudad de México, Ciudad de México 09620, Mexico. edgar.sierra@uacm.edu.mx.
authors.forename: Edgar
authors.initials: E
authors.lastname: Sierra-Palacios
authors.affiliation: Laboratorio de Bioquímica Genética, Instituto Nacional de Pediatría, Secretaría de Salud, Ciudad de México 04530, Mexico. acastilloinp@gmail.com.
authors.forename: Adriana
authors.initials: A
authors.lastname: Castillo-Villanueva
authors.affiliation: CONACYT-Instituto Nacional de Pediatría, Secretaría de Salud, Ciudad de México 04530, Mexico. racastilloro@conacyt.mx.
authors.forename: Rosa Angélica
authors.initials: RA
authors.lastname: Castillo-Rodríguez
authors.affiliation: Departamento de Química de Biomacromoléculas, Instituto de Química, Universidad Nacional Autónoma de México, Circuito Exterior s/n, Ciudad Universitaria, Ciudad de México 04510, Mexico. e-rodriguez-bustamante@ciencias.unam.mx.
authors.forename: Eduardo
authors.initials: E
authors.lastname: Rodríguez-Bustamante
authors.affiliation: Departamento de Química de Biomacromoléculas, Instituto de Química, Universidad Nacional Autónoma de México, Circuito Exterior s/n, Ciudad Universitaria, Ciudad de México 04510, Mexico. arrespin@unam.mx.
authors.forename: Roberto
authors.initials: R
authors.lastname: Arreguin-Espinosa
authors.affiliation: Laboratorio de Bioquímica Genética, Instituto Nacional de Pediatría, Secretaría de Salud, Ciudad de México 04530, Mexico. hreyesvivas@yahoo.com.mx.
authors.forename: Horacio
authors.initials: H
authors.lastname: Reyes-Vivas
keywords: Giardia lamblia; microRNA; small nucleolar RNA; stem-loop RT-qPCR