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Development of A PCR - Cycle Sequencing System for Animal Mitochondrial DNA Analysis

A system for the study of sequence polymorphisms in animal mitochondrial DNA was developed. This system consists of two candidate markers which are mitochondrial cytochrome b and D-loop. Six novel primers were constructed for their amplifications by polymerase chain reaction (PCR). The primers K...

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Main Author: Lau, Chin Hoon
Format: Thesis
Language:English
English
Published: 1994
Online Access:http://psasir.upm.edu.my/id/eprint/8591/1/FSAS_1994_10_A.pdf
http://psasir.upm.edu.my/id/eprint/8591/
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spelling my.upm.eprints.85912011-08-19T04:15:19Z http://psasir.upm.edu.my/id/eprint/8591/ Development of A PCR - Cycle Sequencing System for Animal Mitochondrial DNA Analysis Lau, Chin Hoon A system for the study of sequence polymorphisms in animal mitochondrial DNA was developed. This system consists of two candidate markers which are mitochondrial cytochrome b and D-loop. Six novel primers were constructed for their amplifications by polymerase chain reaction (PCR). The primers KY1 and KY2 amplified a 359-bp port ion of the cytochrome b gene from chicken, water buffalo, horse, goat and sheep. Primers UPM257 and UPM258 were used to synthesize a 1.4 Kb-fragment containing the complete chicken mi tochondrial D-loop. Primers LCH4-DO and LCH2-UP paired with primers UPM257 and UPM258 respectively to amplify a 355-bp 5' region and 395-bp 3' region of the D-loop. These primers were specific in their amplifications. The mitochondrial DNA could be amplified from heterogeneous or degraded DNA preparations, as well as directly from blood incubated in proteinase K thus bypassing lengthy DNA isolation steps.Only 5ul of blood was required for a l00-ul PCR. Fort he purification of PCR products, ethanol selective precipitation was found to be both efficient and economical. Despite its lower recovery compared to the Chroma Spin column, sufficient quantity of DNA could be obtained for subsequent sequencing. 1994 Thesis NonPeerReviewed application/pdf en http://psasir.upm.edu.my/id/eprint/8591/1/FSAS_1994_10_A.pdf Lau, Chin Hoon (1994) Development of A PCR - Cycle Sequencing System for Animal Mitochondrial DNA Analysis. Masters thesis, Universiti Pertanian Malaysia. English
institution Universiti Putra Malaysia
building UPM Library
collection Institutional Repository
continent Asia
country Malaysia
content_provider Universiti Putra Malaysia
content_source UPM Institutional Repository
url_provider http://psasir.upm.edu.my/
language English
English
description A system for the study of sequence polymorphisms in animal mitochondrial DNA was developed. This system consists of two candidate markers which are mitochondrial cytochrome b and D-loop. Six novel primers were constructed for their amplifications by polymerase chain reaction (PCR). The primers KY1 and KY2 amplified a 359-bp port ion of the cytochrome b gene from chicken, water buffalo, horse, goat and sheep. Primers UPM257 and UPM258 were used to synthesize a 1.4 Kb-fragment containing the complete chicken mi tochondrial D-loop. Primers LCH4-DO and LCH2-UP paired with primers UPM257 and UPM258 respectively to amplify a 355-bp 5' region and 395-bp 3' region of the D-loop. These primers were specific in their amplifications. The mitochondrial DNA could be amplified from heterogeneous or degraded DNA preparations, as well as directly from blood incubated in proteinase K thus bypassing lengthy DNA isolation steps.Only 5ul of blood was required for a l00-ul PCR. Fort he purification of PCR products, ethanol selective precipitation was found to be both efficient and economical. Despite its lower recovery compared to the Chroma Spin column, sufficient quantity of DNA could be obtained for subsequent sequencing.
format Thesis
author Lau, Chin Hoon
spellingShingle Lau, Chin Hoon
Development of A PCR - Cycle Sequencing System for Animal Mitochondrial DNA Analysis
author_facet Lau, Chin Hoon
author_sort Lau, Chin Hoon
title Development of A PCR - Cycle Sequencing System for Animal Mitochondrial DNA Analysis
title_short Development of A PCR - Cycle Sequencing System for Animal Mitochondrial DNA Analysis
title_full Development of A PCR - Cycle Sequencing System for Animal Mitochondrial DNA Analysis
title_fullStr Development of A PCR - Cycle Sequencing System for Animal Mitochondrial DNA Analysis
title_full_unstemmed Development of A PCR - Cycle Sequencing System for Animal Mitochondrial DNA Analysis
title_sort development of a pcr - cycle sequencing system for animal mitochondrial dna analysis
publishDate 1994
url http://psasir.upm.edu.my/id/eprint/8591/1/FSAS_1994_10_A.pdf
http://psasir.upm.edu.my/id/eprint/8591/
_version_ 1643824061717938176
score 13.211869

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