Band broadening of DNA fragments isolated by polyacrylamide gel electrophoresis in capillary electrophoresis

Takashi Kaneta, Takehito Ogura, Shuhei Yamato, Totaro Imasaka

Research output: Contribution to journalArticle

12 Citations (Scopus)

Abstract

Polyacrylamide gel electrophoresis (PAGE) is used frequently for isolation and purification of DNA fragments. In the present study, DNA fragments extracted from polyacrylamide gels showed significant band broadening in capillary electrophoresis (CE). A pHY300PLK (a shuttle vector functioning in Escherichia coli and Bacillus subtilis) marker, which contained nine fragments ranging from 80 to 4870 bp, was separated by PAGE, and each fragment was isolated by phenol/chloroform extraction and ethanol precipitation. After extraction from the polyacrylamide gel, the peaks of the isolated DNA fragments exhibited band broadening in CE, where a linear poly(ethylene oxide) was used as a sieving matrix. The theoretical plate numbers of the DNA fragments contained in the pHY300PLK marker were >10 6 for all the fragments before extraction. However, the DNA fragments extracted from the polyacrylamide gel showed decreased theoretical plate numbers (5-20 times smaller). The degradation of the theoretical plate number was significant for middle sizes of the DNA fragments ranging from 489 to 1360 bp, whereas the largest and smallest fragments (80 and 4870 bp) had no obvious influence. The band broadening was attributed to contamination of the DNA fragments by polyacrylamide fibers during the separation and extraction process.

Original languageEnglish
Pages (from-to)431-435
Number of pages5
JournalJournal of Separation Science
Volume35
Issue number3
DOIs
Publication statusPublished - Feb 2012

Fingerprint

Capillary electrophoresis
Polyacrylates
Electrophoresis
DNA
Gels
Genetic Vectors
Bacilli
Chloroform
Chlorine compounds
Polyethylene oxides
Phenol
polyacrylamide gels
Escherichia coli
Phenols
Purification
Contamination
Ethanol
Degradation
Fibers

Keywords

  • Band broadening
  • Capillary electrophoresis
  • DNA
  • Polyacrylamide gel electrophoresis

ASJC Scopus subject areas

  • Analytical Chemistry
  • Filtration and Separation

Cite this

Band broadening of DNA fragments isolated by polyacrylamide gel electrophoresis in capillary electrophoresis. / Kaneta, Takashi; Ogura, Takehito; Yamato, Shuhei; Imasaka, Totaro.

In: Journal of Separation Science, Vol. 35, No. 3, 02.2012, p. 431-435.

Research output: Contribution to journalArticle

@article{a566900912634a10b0a5e407fe8c1c90,
title = "Band broadening of DNA fragments isolated by polyacrylamide gel electrophoresis in capillary electrophoresis",
abstract = "Polyacrylamide gel electrophoresis (PAGE) is used frequently for isolation and purification of DNA fragments. In the present study, DNA fragments extracted from polyacrylamide gels showed significant band broadening in capillary electrophoresis (CE). A pHY300PLK (a shuttle vector functioning in Escherichia coli and Bacillus subtilis) marker, which contained nine fragments ranging from 80 to 4870 bp, was separated by PAGE, and each fragment was isolated by phenol/chloroform extraction and ethanol precipitation. After extraction from the polyacrylamide gel, the peaks of the isolated DNA fragments exhibited band broadening in CE, where a linear poly(ethylene oxide) was used as a sieving matrix. The theoretical plate numbers of the DNA fragments contained in the pHY300PLK marker were >10 6 for all the fragments before extraction. However, the DNA fragments extracted from the polyacrylamide gel showed decreased theoretical plate numbers (5-20 times smaller). The degradation of the theoretical plate number was significant for middle sizes of the DNA fragments ranging from 489 to 1360 bp, whereas the largest and smallest fragments (80 and 4870 bp) had no obvious influence. The band broadening was attributed to contamination of the DNA fragments by polyacrylamide fibers during the separation and extraction process.",
keywords = "Band broadening, Capillary electrophoresis, DNA, Polyacrylamide gel electrophoresis",
author = "Takashi Kaneta and Takehito Ogura and Shuhei Yamato and Totaro Imasaka",
year = "2012",
month = "2",
doi = "10.1002/jssc.201100909",
language = "English",
volume = "35",
pages = "431--435",
journal = "Journal of Separation Science",
issn = "1615-9306",
publisher = "Wiley-VCH Verlag",
number = "3",

}

TY - JOUR

T1 - Band broadening of DNA fragments isolated by polyacrylamide gel electrophoresis in capillary electrophoresis

AU - Kaneta, Takashi

AU - Ogura, Takehito

AU - Yamato, Shuhei

AU - Imasaka, Totaro

PY - 2012/2

Y1 - 2012/2

N2 - Polyacrylamide gel electrophoresis (PAGE) is used frequently for isolation and purification of DNA fragments. In the present study, DNA fragments extracted from polyacrylamide gels showed significant band broadening in capillary electrophoresis (CE). A pHY300PLK (a shuttle vector functioning in Escherichia coli and Bacillus subtilis) marker, which contained nine fragments ranging from 80 to 4870 bp, was separated by PAGE, and each fragment was isolated by phenol/chloroform extraction and ethanol precipitation. After extraction from the polyacrylamide gel, the peaks of the isolated DNA fragments exhibited band broadening in CE, where a linear poly(ethylene oxide) was used as a sieving matrix. The theoretical plate numbers of the DNA fragments contained in the pHY300PLK marker were >10 6 for all the fragments before extraction. However, the DNA fragments extracted from the polyacrylamide gel showed decreased theoretical plate numbers (5-20 times smaller). The degradation of the theoretical plate number was significant for middle sizes of the DNA fragments ranging from 489 to 1360 bp, whereas the largest and smallest fragments (80 and 4870 bp) had no obvious influence. The band broadening was attributed to contamination of the DNA fragments by polyacrylamide fibers during the separation and extraction process.

AB - Polyacrylamide gel electrophoresis (PAGE) is used frequently for isolation and purification of DNA fragments. In the present study, DNA fragments extracted from polyacrylamide gels showed significant band broadening in capillary electrophoresis (CE). A pHY300PLK (a shuttle vector functioning in Escherichia coli and Bacillus subtilis) marker, which contained nine fragments ranging from 80 to 4870 bp, was separated by PAGE, and each fragment was isolated by phenol/chloroform extraction and ethanol precipitation. After extraction from the polyacrylamide gel, the peaks of the isolated DNA fragments exhibited band broadening in CE, where a linear poly(ethylene oxide) was used as a sieving matrix. The theoretical plate numbers of the DNA fragments contained in the pHY300PLK marker were >10 6 for all the fragments before extraction. However, the DNA fragments extracted from the polyacrylamide gel showed decreased theoretical plate numbers (5-20 times smaller). The degradation of the theoretical plate number was significant for middle sizes of the DNA fragments ranging from 489 to 1360 bp, whereas the largest and smallest fragments (80 and 4870 bp) had no obvious influence. The band broadening was attributed to contamination of the DNA fragments by polyacrylamide fibers during the separation and extraction process.

KW - Band broadening

KW - Capillary electrophoresis

KW - DNA

KW - Polyacrylamide gel electrophoresis

UR - http://www.scopus.com/inward/record.url?scp=84856085856&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=84856085856&partnerID=8YFLogxK

U2 - 10.1002/jssc.201100909

DO - 10.1002/jssc.201100909

M3 - Article

C2 - 22258810

AN - SCOPUS:84856085856

VL - 35

SP - 431

EP - 435

JO - Journal of Separation Science

JF - Journal of Separation Science

SN - 1615-9306

IS - 3

ER -