Improved procedure for the preparation of DNA restriction fragments suitable for sequencing

Piergiorgio Pletta, Pierluigi Mauri, Valentina Appierto, Marina Mostardini, Ida Biunno

Research output: Contribution to journalArticlepeer-review

Abstract

A rapid and integrated procedure was developed for the preparation of small DNA restriction fragments ( ≤ 1000 bp) starting from a large cosmid (35,000 bp) containing exogenous DNA. The process is based on restriction enzymatic digestion followed by HPLC separation and fractions collection. All DNA fragments are separated in a single run, detected "on-line" by UV absorption, and straightforward collected with very high recovery. Small fragments can be directly subjected to the sequence procedure, whereas those larger than 1000 bp are redigested with a second enzyme, the fractionated subfragments are separated, ligated to plas-mid vector, and sequenced. A human genomic cosmid of 35,000 bp (26H7) has been chosen as a model.

Original languageEnglish
Pages (from-to)119-124
Number of pages6
JournalApplied Biochemistry and Biotechnology
Volume44
Issue number2
DOIs
Publication statusPublished - Feb 1994

Keywords

  • DNA restriction fragments
  • HPLC
  • preparation of

ASJC Scopus subject areas

  • Bioengineering
  • Biotechnology
  • Biochemistry
  • Biochemistry, Genetics and Molecular Biology(all)

Fingerprint

Dive into the research topics of 'Improved procedure for the preparation of DNA restriction fragments suitable for sequencing'. Together they form a unique fingerprint.

Cite this