Dataset Information

Two methods for genotyping a 4-base deletion in the canine ABCB1 gene.

ABSTRACT: A 4-bp deletion (c.230_233delATAG) of the ABCB1 gene, frequently found in various dog breeds, results in intolerance to certain drugs routinely used in veterinary medicine, including many chemotherapeutic agents and macrocyclic lactones. The use of rapid and reliable genetic testing is fundamental for early detection of the mutation and prevention of undesirable toxicoses. We developed and compared 2 genotyping tests: PCR-high-resolution melting (PCR-HRM) and PCR-restriction-fragment length polymorphism (PCR-RFLP) to identify the 4-bp deletion in the ABCB1 gene of canine breeds. Amplified PCR products were sequenced in order to confirm different genotypes. Both techniques were efficient in discriminating homozygous wild-type, homozygous mutated, and heterozygous ABCB1 genotypes, and proved to be reproducible and economical methods. The HRM analysis, a sensitive and specific method for the molecular detection of genetic disorders, does not require labeled probes, processing, or separations after PCR.

SUBMITTER: Silvestro CA

PROVIDER: S-EPMC6900729 | biostudies-literature | 2019 Nov

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Two methods for genotyping a 4-base deletion in the canine <i>ABCB1</i> gene.

Silvestro Carolina A CA Soria Liliana A LA Conte Adriana A Marrube Graciela G

Journal of veterinary diagnostic investigation : official publication of the American Association of Veterinary Laboratory Diagnosticians, Inc 20191101 6

A 4-bp deletion (c.230_233delATAG) of the <i>ABCB1</i> gene, frequently found in various dog breeds, results in intolerance to certain drugs routinely used in veterinary medicine, including many chemotherapeutic agents and macrocyclic lactones. The use of rapid and reliable genetic testing is fundamental for early detection of the mutation and prevention of undesirable toxicoses. We developed and compared 2 genotyping tests: PCR-high-resolution melting (PCR-HRM) and PCR-restriction-fragment leng ...[more]

PMID: 31711409

Similar Datasets

Project description:We compared two consensus primer PCR human papillomavirus (HPV) genotyping methods for the detection of individual HPV genotypes and carcinogenic HPV genotypes as a group, using a stratified sample of enrollment cervical specimens from sexually active women participating in the NCI/Costa Rica HPV16/18 Vaccine Efficacy Trial. For the SPF(10) method, DNA was extracted from 0.1% of the cervical specimen by using a MagNA Pure LC instrument, a 65-bp region of the HPV L1 gene was targeted for PCR amplification by using SPF(10) primers, and 25 genotypes were detected by reverse-line blot hybridization of the amplicons. For the Linear Array (LA) method, DNA was extracted from 0.5% of the cervical specimen by using an MDx robot, a 450-bp region of the HPV L1 gene was targeted for PCR amplification by using PGMY09/11 L1 primers, and 37 genotypes were detected by reverse-line blot hybridization of the amplicons. Specimens (n = 1,427) for testing by the LA method were randomly selected from strata defined on the basis of enrollment test results from the SPF(10) method, cytology, and Hybrid Capture 2. LA results were extrapolated to the trial cohort (n = 5,659). The LA and SPF(10) methods detected 21 genotypes in common; HPV16, -18, -31, -33, -35, -39, -45, -51, -52, -56, -58, -59, -66, -68, and -73 were considered the carcinogenic HPV genotypes. There was no difference in the overall results for grouped detection of carcinogenic HPV by the SPF(10) and LA methods (35.3% versus 35.9%, respectively; P = 0.5), with a 91.8% overall agreement and a kappa value of 0.82. In comparisons of individual HPV genotypes, the LA method detected significantly more HPV16, HPV18, HPV39, HPV58, HPV59, HPV66, and HPV68/73 and less HPV31 and HPV52 than the SPF(10) method; inclusion of genotype-specific testing for HPV16 and HPV18 for those specimens testing positive for HPV by the SPF(10) method but for which no individual HPV genotype was detected abrogated any differences between the LA and SPF(10) methods. The LA method detected more carcinogenic-HPV-genotype infections per specimen than the SPF(10) method (P < 0.001). In conclusion, the LA method and the SPF(10) method with HPV16 and HPV18 genotype-specific detection among ungenotyped HPV-positive specimens were comparable for detection of HPV16 and HPV18, the two HPV genotypes targeted by current prophylactic HPV vaccines. Both approaches are suitable for monitoring the impact of HPV16/18 vaccines in clinical trials.

Project description:BACKGROUND:Since most prostatic diseases are associated with the organ's enlargement, evaluation of prostatic size is a main criterion in the diagnosis of prostatic state of health. While enlargement is a non-uniform process, volumetric measurements are believed to be advantageous to any single dimensional parameter for the diagnosis of prostatomegaly. In a previous study, volume was analysed with a slice addition technique (SAT), which was validated as highly accurate. Irrespective of high accuracy, SAT represents a complex and time-consuming procedure, which limits its clinical use. Thus, demand exists for more practical volume assessment methods. In this study, the prostatic volume of 95 canine patients (58 intact males, 37 neutered males) were analysed retrospectively by using the ellipsoid formula (Formula) and an imaging "wrap" function tool (Wrap) to help assess accuracy and applicability. Accuracy was checked against phantom measurements and results were compared to SAT measurements of the same patient pool obtained from a previously published paper. Patients were grouped according to prostatic structure (H?=?homogeneous, I?=?inhomogeneous, C?=?cystic) and volume using the SAT (volume group?=?vg: 1, 2 and 3). RESULTS:High correlation between the Formula or Wrap volume and the phantom volume was found, the values being higher for the Formula. Mean Formula volumes (vg 1: 2.2?cm3, vg 2: 14.5?cm3, vg 3: 109.4?cm3, respectively) were significantly underestimated, while mean Wrap volumes (vg 1: 3.8?cm3, vg 2: 19.5?cm3, vg 3: 159.2?cm3) were statistically equivalent to SAT measurements (vg 1: 3.1?cm3, vg 2: 18.6?cm3, vg 3: 157.2?cm3, respectively). Differences between Formula and SAT volumes ranged from 22.4-31.1%, while differences between Wrap and SAT volumes were highest in small prostates (vg 1: 22.1%) and fell with increasing prostatic size (vg 3: 1.3%). CONCLUSION:The Wrap function is highly accurate, less time-consuming and complex compared to SAT and could serve as beneficial tool for measuring prostatic volume in clinical routine after further validation in future studies. The Formula method cannot be recommended as an alternative for volumetric measurements of the prostate gland due to its underestimation of volumes compared to SAT results.

Dataset Information

Two methods for genotyping a 4-base deletion in the canine ABCB1 gene.

Publications

Two methods for genotyping a 4-base deletion in the canine <i>ABCB1</i> gene.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets