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SYBR® Green quantitative PCR for sex determination of bovine spermatozoa

Spermatozoa sexing technology in cattle breeding is being done by separation of X- and Y- chromosome bearing spermatozoa using flow-sorting technology. However, the sexing technique needed to be validated to ensure the accuracy of the technology. A technique to determine the sex of bovine spermatozo...

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Main Authors: Tan, Y. J., Mahanem Mat Noor, Wan Somarny Wan Md. Zain
Format: Journal Contribution
Language:English
Published: 2020
Subjects:
Bovines
Spermatozoa
PCR
Sex determination
Sexing
Cattle
Animal breeding
Chromosomes
Separation
High temperature
Plasmids
Online Access:http://agris.upm.edu.my:8080/dspace/handle/0/17448
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spelling oai:http:--agris.upm.edu.my:0-17448SYBR® Green quantitative PCR for sex determination of bovine spermatozoaTan, Y. J.Mahanem Mat NoorWan Somarny Wan Md. ZainBovinesSpermatozoaPCRSex determinationSexingCattleAnimal breedingChromosomesSeparationHigh temperaturePlasmidsSpermatozoa sexing technology in cattle breeding is being done by separation of X- and Y- chromosome bearing spermatozoa using flow-sorting technology. However, the sexing technique needed to be validated to ensure the accuracy of the technology. A technique to determine the sex of bovine spermatozoa using SYBR® Green real-time quantitative PCR (qPCR) was developed. Two sets of primers, ZFX and SRY were designed specifically to X- and Y- chromosome bovine genes respectively. Plasmid was inserted with ZFX and SRY gene fragment separately to create standard curves that ranged from 3.0 x 102 to 3.0 x 106 copies. The standards generated linear relationship with regression coefficient r2 = 0.984 for ZFX and r2 = 0.996 for SRY. Both standards of ZFX and SRY showed melting peak at temperature of 83 ºC and 85 ºC respectively. Real-time qPCR of bovine spermatozoa DNA samples and cloned plasmid ZFX and SRY genes for creating standard samples were performed simultaneously. The percentages of unsexed X- and Y- chromosome-bearing spermatozoa did not differ much from the 1:1 as reported in unsexed spermatozoa population. Therefore, the highly sensitive and fast method of real time PCR is a suitable technique for quantitating X- and Y- chromosome-bearing spermatozoa in semen samples.2020-07-07T08:10:31Z2020-07-07T08:10:31Z2015Journal ContributionArticleRefereedJournal of Tropical Agriculture and Food Science (Malaysia), 43 (1), p. 29-391394-9829http://agris.upm.edu.my:8080/dspace/handle/0/17448MY2020050754enhttp://jtafs.mardi.gov.my/jtafs/43-1/SYBR.pdfMalaysiahttp://www.oceandocs.org/license
institution AGRIS
collection AGRIS
language English
topic Bovines
Spermatozoa
PCR
Sex determination
Sexing
Cattle
Animal breeding
Chromosomes
Separation
High temperature
Plasmids
spellingShingle Bovines
Spermatozoa
PCR
Sex determination
Sexing
Cattle
Animal breeding
Chromosomes
Separation
High temperature
Plasmids
Tan, Y. J.
Mahanem Mat Noor
Wan Somarny Wan Md. Zain
SYBR® Green quantitative PCR for sex determination of bovine spermatozoa
description Spermatozoa sexing technology in cattle breeding is being done by separation of X- and Y- chromosome bearing spermatozoa using flow-sorting technology. However, the sexing technique needed to be validated to ensure the accuracy of the technology. A technique to determine the sex of bovine spermatozoa using SYBR® Green real-time quantitative PCR (qPCR) was developed. Two sets of primers, ZFX and SRY were designed specifically to X- and Y- chromosome bovine genes respectively. Plasmid was inserted with ZFX and SRY gene fragment separately to create standard curves that ranged from 3.0 x 102 to 3.0 x 106 copies. The standards generated linear relationship with regression coefficient r2 = 0.984 for ZFX and r2 = 0.996 for SRY. Both standards of ZFX and SRY showed melting peak at temperature of 83 ºC and 85 ºC respectively. Real-time qPCR of bovine spermatozoa DNA samples and cloned plasmid ZFX and SRY genes for creating standard samples were performed simultaneously. The percentages of unsexed X- and Y- chromosome-bearing spermatozoa did not differ much from the 1:1 as reported in unsexed spermatozoa population. Therefore, the highly sensitive and fast method of real time PCR is a suitable technique for quantitating X- and Y- chromosome-bearing spermatozoa in semen samples.
format Journal Contribution
author Tan, Y. J.
Mahanem Mat Noor
Wan Somarny Wan Md. Zain
author_facet Tan, Y. J.
Mahanem Mat Noor
Wan Somarny Wan Md. Zain
author_sort Tan, Y. J.
title SYBR® Green quantitative PCR for sex determination of bovine spermatozoa
title_short SYBR® Green quantitative PCR for sex determination of bovine spermatozoa
title_full SYBR® Green quantitative PCR for sex determination of bovine spermatozoa
title_fullStr SYBR® Green quantitative PCR for sex determination of bovine spermatozoa
title_full_unstemmed SYBR® Green quantitative PCR for sex determination of bovine spermatozoa
title_sort sybr® green quantitative pcr for sex determination of bovine spermatozoa
publishDate 2020
url http://agris.upm.edu.my:8080/dspace/handle/0/17448
_version_ 1819285313006075904
score 13.4562235

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