Potato blight causes potato plants to shrivel and rot. The disease is caused by the pathogen Phytophthora infestans, infamous for its role in Ireland's Great Potato Famine in the mid-1840s. The disease can devastate crops during wet weather, sometimes leading to total crop loss. Researchers aim to use recombinant DNA methods to transfer blight resistance genes from resistant varieties into susceptible varieties of potato. Explain how restriction endonucleases and DNA ligase could be used to insert a potato blight resistance gene into a plasmid.

Verified step by step guidance

1. Identify and isolate the gene responsible for blight resistance in a potato variety that is naturally resistant to Phytophthora infestans. This gene will be the segment of DNA that needs to be inserted into the susceptible potato variety.
2. Use restriction endonucleases to cut the DNA at specific recognition sites. These enzymes act as molecular scissors to cut both the DNA containing the blight resistance gene and the plasmid DNA at specific sequences, creating sticky ends or blunt ends.
3. Mix the cut plasmid DNA with the DNA fragment containing the blight resistance gene. The sticky ends of the plasmid DNA and the gene should complement each other, allowing them to anneal or join together through base pairing.
4. Add DNA ligase to the mixture. DNA ligase is an enzyme that facilitates the formation of phosphodiester bonds between adjacent nucleotides, effectively sealing the nicks in the sugar-phosphate backbone of the DNA. This step permanently integrates the blight resistance gene into the plasmid.
5. Introduce the recombinant plasmid into susceptible potato cells through a process such as transformation, where cells take up the plasmid from their environment. The cells can then be grown to develop into plants that express the blight resistance gene, thereby inheriting resistance to potato blight.

Key Concepts

Restriction Endonucleases

Restriction endonucleases, or restriction enzymes, are proteins that cut DNA at specific sequences, creating fragments with 'sticky' or 'blunt' ends. These enzymes are crucial in genetic engineering as they allow scientists to isolate specific genes, such as those conferring resistance to potato blight. By cutting both the plasmid and the target gene with the same restriction enzyme, compatible ends are generated, facilitating the insertion of the gene into the plasmid.

DNA Ligase

DNA ligase is an enzyme that facilitates the joining of DNA strands together by forming phosphodiester bonds. In the context of genetic engineering, after a gene of interest has been inserted into a plasmid, DNA ligase is used to seal the nicks in the sugar-phosphate backbone, ensuring that the plasmid is a continuous, stable piece of DNA. This step is essential for the successful incorporation of the resistance gene into the plasmid, allowing for its propagation in host cells.

Recombinant DNA Technology

Recombinant DNA technology involves combining DNA from different sources to create new genetic combinations. This technique is pivotal in transferring desirable traits, such as disease resistance, into crops. By using restriction endonucleases to cut DNA and DNA ligase to join it, researchers can create plasmids that carry the blight resistance gene, which can then be introduced into susceptible potato varieties, enhancing their resilience against diseases like potato blight.