8.5.3 Gene Cloning and Transformation

• Gene cloning is the process of making multiple identical copies of a gene.
• The transformation step in gene cloning involves introducing the recombinant plasmid into host cells, typically bacteria.


  True
• What is the purpose of cloning vectors in gene cloning?

  To carry the target gene
• The origin of replication in a cloning vector allows the vector to replicate independently within the host cell.
• Marker genes such as antibiotic resistance are used to identify successful transformants.


  True
• Which enzyme is used to extract the target gene from its source DNA?

  Restriction enzymes
• Plasmids are cloning vectors capable of carrying up to 15,000 base pairs.
• Plasmids can carry DNA fragments up to 15,000 base pairs in size.


  True
• Match the cloning vector type with its capacity (base pairs):

  Plasmids ↔️ Up to 15,000

  Cosmids ↔️ Up to 45,000

  BACs ↔️ 100,000 to 300,000

  YACs ↔️ Up to 2,000,000
• Which type of cloning vector is used for cloning extremely large DNA segments in yeast?

  YACs
• The origin of replication in a cloning vector allows it to replicate independently within the host cell.


  True
• Multiple cloning sites in a cloning vector are regions with restriction enzyme sites for inserting the target gene.
• What are the two main methods used in host cell transformation?

  Heat shock and electroporation
• Steps in host cell transformation using heat shock

  1️⃣ Host cells are mixed with the plasmid

  2️⃣ Cells are rapidly heated

  3️⃣ Pores form in the cell membrane

  4️⃣ Plasmid enters the cell
• Electroporation uses electrical pulses to create temporary pores in the cell membrane.

  True
• Successful transformants are selected using marker genes such as antibiotic resistance.
• What is one application of gene cloning in the field of medicine?

  Protein production
• What enzyme is used to insert the target gene into a plasmid vector?

  Ligase
• The desired gene is extracted from source DNA using restriction enzymes.
• Match the cloning vector type with its capacity and common uses:

  Plasmids ↔️ Up to 15,000 base pairs; simple gene cloning in bacteria

  Phages ↔️ Up to 25,000 base pairs; cloning larger fragments in bacteria

  BACs ↔️ 100,000 to 300,000 base pairs; cloning very large DNA fragments in bacteria
• What are two common methods used in host cell transformation?

  Heat shock and electroporation
• Gene cloning involves inserting a target gene into a cloning vector.
• Steps in the gene cloning process:

  1️⃣ Isolation of the target gene

  2️⃣ Insertion into a vector

  3️⃣ Transformation into host cells

  4️⃣ Multiplication and selection
• Which cloning vector is capable of cloning extremely large DNA segments in yeast?

  YACs
• The origin of replication in a cloning vector allows it to replicate independently within the host cell.


  True
• What are cloning vectors used for in the gene cloning process?

  Carrying a target gene
• BACs are capable of cloning DNA fragments ranging from 100,000 to 300,000 base pairs.
• Cloning can be used in gene therapy to produce therapeutic genes for treating genetic disorders.


  True