Various Stages of Plasmid Isolation: A Part from The Book Chapter : Homemade Molecular Devices, Detections and Methods

DNA degrading enzymes

Often the DNase-1 is thermostable and it is not possible for its inactivation by standard or enhanced thermal treatment. It is reported that heat treatment (at various stages of plasmid isolation) for inactivating the nucleases may reduce the chromosomal smearing, but did not improve plasmid recovery. The enzyme DNase has been reported as thermo-resistant and responsible for reduced DNA recovery. Although heat partially inactivated the DNase present in each cell fraction, the reduced inhibition suggest that DNase of C. pasteurianum was resistant to thermal inactivation. This affects the use and preservation of DNA.

These enzymes however, are known to be dependent on cofactors like Mg+2, Mn+2 or Ca+2 for their action. These cofactors can be chelated by sodium citrate, EDTA etc. thus inhibiting the enzymes that cause degradation of DNA. Chelation does not affect DNA isolation nor its preservation but inhibit only the DNA degrading enzymes.

Author(s) Details:

Lahiri S. S.,
Defence R & D Est., D.R.D.O., Gwalior, Madhya Pradesh, India and Institute of Nuclear Medicine and Allied Science (INMAS), DRDO, New Delhi, India and Amity University, Noida, Uttar Pradesh, India and Amity University, Noida, Uttar Pradesh, India.

Singh R.,
Defence R & D Est., D.R.D.O., Gwalior, Madhya Pradesh, India.

Karothia B. S.,
Defence R & D Est., D.R.D.O., Gwalior, Madhya Pradesh, India.

Giredhar M.,
Center for Bio-Hybrid Medical Systems, Forckenbeckstr. 55, 52074 Aachen, Germany.

Chakraborty A.,
Division of Immunology, Immunity to Infection and Respiratory Medicine, School of Biological Sciences, The University of Manchester, Manchester, M13 9PT, UK.

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