Improved Long ssDNA Preparation Kits

news December 23 2019

Long single strand DNA (ssDNA) preparation kit – for effective knock-in by genome editing!

The Long ssDNA Preparation Kits (LssDNA Preparation Kits) provide a simple and easy method for generating a long ssDNA (<1,500 base or< 3,000 base). A long ssDNA prepared by this kit has a defined sequence and length as it does not include internal mutation and terminal deletion caused by PCR, exonuclease side reaction, low-fidelity reverse transcriptase reaction or low-fidelity synthetic oligonucleotides.

The procedure is almost the same as the method to obtain dsDNA fragments. The DNA of interest is cloned into a plasmid. The resulting plasmid harbouring the DNA is digested by a pair of two nicking endonucleases or a combination of a nicking endonuclease and a restriction enzyme. The nicked plasmid is denatured by Denaturing Gel-Loading Buffer and then subjected to agarose gel electrophoresis. The band corresponding to a long ssDNA is excised and extracted with our newly developed extraction kits.

Features

  • A long ssDNA (< 1,500 base or < 3,000 base) can be prepared.
  • A long ssDNA has a defined sequence and length.
  • Simple principle and easy procedure.
  • High yield and high quality.

Principle

(1) The DNA of interest is cloned into a plasmid using a pair of two nicking endonuclease sites or a combination of a nicking endonuclease site and a restriction enzyme site. (2) The resulting plasmid harbouring the DNA is digested with a pair of two nicking endonucleases or a combination of a nicking endonuclease and a restriction enzyme. (3) The nicked plasmid is denatured and then subjected to agarose gel electrophoresis. (4) The band corresponding to a long ssDNA is excised and extracted.

Plasmid Map

Data

Fig.1 Agarose gel electrophoresis of purified long ssDNAs. 1.2% agarose gel in 1x TAE. 5.5cm x 6.0 cm. Lane 1: pLSODN-1 (1.5 kb Fragment), double-nicked and denatured 800 ng Lane 2: a long ssDNA purified from pLSODN-1 (1.5 kb Fragment) 400 ng Lane 3: pLSODN-3 (3 kb Fragment), double-nicked and denatured 800 ng Lane 4: a long ssDNA purified from pLSODN-3 (3 kb Fragment) 400 ng

Kit Contents

DS615 Long ssDNA Preparation Kit for 1.5 kb (LsODN Preparation Kit)

  • pLSODN-1 10 μg (0.5 μg/μl)
  • pLSODN-2D 10 μg (0.5 μg/μl)
  • Denaturing Gel-Loading Buffer 1 ml (100 loadings)
  • Nicked pLSODN-1 (1.5 kb Fragment)
  • Long ssDNA Gel Extraction Kit for 3kb (25 preps)

DS625 Long ssDNA Preparation Kit for 3.0 kb (LsODN Preparation Kit)

  • pLSODN-3 10 μg (0.5 μg/μl )
  • pLSODN-4D 10 μg (0.5 μg/μl )
  • Denaturing Gel-Loading Buffer 1 ml (100 loadings)
  • Nicked pLSODN-3 (3.0 kb Fragment)
  • Long ssDNA Gel Extraction Kit for 3kb (25 preps)