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MiniPCR Chopped! CRISPR Activity - Student Guide

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miniPCR bio TM Chopped! Using CRISPR/Cas9 to cut DNA - Student's Guide Version: 1.0 - Release: May 2022 - © 2022 by miniPCR bio™ Student's Guide P./12 Background information Overview - The introduction of CRISPR/Cas9 into the biotechnology toolkit has revolutionized scientists' ability to manipulate DNA in living organisms. In today's lab, you will use the CRISPR/Cas9 system in vitro (in a test tube) to break down exactly how the different parts of this gene targeting system work. CRISPR/Cas9: a powerful tool - Typically, an organism's DNA sequence doesn't change over its lifespan. But sometimes scientists want to change the DNA within the cells of a living organism. For example, they may wish to investigate how certain genes work or correct mutations that cause genetic disease. Making specific changes to DNA inside living cells is called genome editing, and developing reliable genome editing tools has been the goal of scientists for decades. Until recently, the methods used to perform genome editing could only be applied to a few model organisms such as mice. Furthermore, these genome editing tools were generally inefficient and difficult to use. That changed in 2012 with the introduction of the CRISPR/Cas9 system. The CRISPR/ Cas9 system provided a universal method to make targeted changes to almost any DNA sequence in virtually any organism. This proved revolutionary both for scientists trying to understand the functions of genes and for clinical researchers seeking to treat genetic diseases. To understand why CRISPR/Cas9 is such a versatile system, we need to look at its individual components and understand how they work at the molecular level. Using CRISPR/Cas9 as a genome editing tool - Scientists did not invent the CRISPR/Cas system from scratch; like most biotechnology tools, it has a natural origin. CRISPR/Cas is found naturally in bacteria and archaea where it plays a role in immune function, providing protection from harmful viruses. Scientists have repurposed this bacterial immune system as a genome editing tool because it allows them to target specific DNA sequences with relative ease. The Cas in CRISPR/Cas refers to an enzyme known as the Cas nuclease. A nuclease is an enzyme that cuts nucleic acids, such as DNA, like a pair of molecular scissors. There are actually several

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