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Chapter 13

Biotechnology

13.1 What Is Biotechnology?

 (^) Biotechnology is the use and alteration, of organisms, cells, or biological molecules to produce food, drugs, or other goods

• (^) use of yeast to make bread, beer, and wine
• (^) selective breeding of animals for desired traits  (^) More modern uses modify the genes directly through genetic engineering
• (^) A key tool is creating recombinant DNA (DNA that has been altered to contain genes or portions of genes from different organisms)
• (^) Plants and animals that express DNA that has been modified or derived from other species are transgenic or genetically modified organisms (GMOs)

What are some of the pros & cons of GMO foods?

13.2 How Does DNA Recombine in Nature?

 Sexual reproduction recombines DNA

• (^) During meiosis I, homologous chromosomes exchange DNA
• (^) Therefore, every egg & every sperm contain recombinant DNA derived from the two parents

 Transformation may combine DNA from different

bacterial species

• (^) In transformation , bacteria pick up pieces of DNA from the environment
• (^) This DNA could be part of a chromosome from another bacterium or tiny circular DNA molecules called plasmids

 What are plasmids?

• (^) Small pieces of DNA, not included in the chromosomes, that may be present in bacteria in multiple copies
• (^) Carry genes that help the bacteria survive in novel environments or where antibiotics are present

Author Animation: Bacterial Transformation

plasmid plasmid bacterial chromosome (a) Bacterium DNA fragments bacterial chromosome (b) Transformation with a DNA fragment (c) Transformation with a plasmid A DNA fragment is incorporated into the chromosome bacterial chromosome The plasmid replicates in the cytoplasm 1 micrometer

Transformation in Bacteria

Fig. 13-

 Viruses may transfer DNA between species

• (^) Viruses can only reproduce inside other living cells, and not alive themselves
• (^) Viral reproduction follows several steps
  1. Attaches to specific molecules on the surface of a suitable host cell
  2. Enters the cytoplasm of the host cell
  3. Virus releases its genetic material
  4. Host replicates the viral genetic material and synthesizes viral proteins
  5. The replicated genes and viral proteins assemble inside the cell
  6. New viruses are formed, which are released and may infect new cells

The Life Cycle of a Typical Virus

Fig. 13- The virus releases its DNA into the host cell; some viral DNA (red) may be incorporated into the host cell’s DNA (blue) The virus enters the host cell Viral genes encode the synthesis of viral proteins and viral gene replication; some host cell DNA may attach to the replicated viral New viruses assemble; DNA (red/blue combination) some host cell DNA is carried by recombinant viruses The host cell bursts open, releasing newly assembled viruses; if recombinant viruses infect a second cell, they may transfer genes from the first cell to the second cell recombinant virus^ viral proteins virus viral DNA viral DNA A virus attaches to a susceptible host cell host cell host cell DNA 2 3 1 4 5 6

Author Animation: Viruses in Genetic

Recombination II

 Viruses may transfer DNA between species ( cont.)

• (^) Most viruses infect specific host species
• (^) Some viruses can infect multiple species
  • (^) Human flu and bird flu epidemics of 1957 and 1968
  • (^) Caused over 100,000 deaths

13.3 How Is Biotechnology Used in Forensic Science?

 The polymerase chain reaction (PCR) amplifies

DNA

• (^) Developed by Kary Mullis of the Cetus Corporation
• (^) Polymerase chain reaction produces virtually unlimited copies of a very small DNA sample
• (^) DNA left behind in tiny amounts can be from crime scenes, murders, fossils, or ancient material
• (^) PCR is used by forensic scientists to identify victims and criminals, and is used extensively in biotechnology and biomedicine

 The polymerase chain reaction amplifies DNA

(contd.)

• (^) Sequence to be copied is heated
• (^) PCR requires a small piece of DNA (called a primer) that is complementary to the gene sequences targeted for copying
• (^) DNA polymerase uses free nucleotides to create complementary strands (very heat STABLE)
• (^) Doubles number of copies of DNA in each cycle
• (^) “Xerox” machine for DNA

 A PCR cycle involves 4 steps

1. Template strand separation - Sample is heated to 90-95°C to cause the double stranded template DNA to separate into single strands… 2. Binding of the primers - The temperature is lowered to 50oC to allow the primer DNA segments to bind to the targeted gene sequences through hydrogen bonding…

 A PCR cycle involves 4 steps (continued

3. New DNA synthesis at targeted sequences - The temperature is raised to 70-72oC where the heat-stable DNA-polymerase synthesizes new DNA of the sequences targeted by the primers… 4. Repetition of the cycle - The cycle is repeated automatically (by a thermocycler machine) for 20-30 cycles, producing up to 1 billion copies of the original targeted DNA sequence

Polymerase Chain

Reaction

Double-stranded DNA to copy DNA heated to 90°– 94°C Primers added to base-pair with ends Mixture cooled; base-pairing of primers and ends of DNA strands DNA polymerases assemble new DNA strands

Mixture heated again; makes all DNA fragments unwind Mixture cooled; base- pairing between primers and ends of single DNA strands DNA polymerase action again doubles number of identical DNA fragments

Polymerase Chain

Reaction

PCR Copies a Specific DNA Sequence

Fig. 13- One PCR cycle original DNA 90 °C 50 °C 72 °C DNA polymerase new DNA strands primers 1 Heating separates DNA strands. 2 Cooling allows primers and DNA polymerase to bind. 3 New DNA strands are synthesized.

PCR Copies a Specific DNA Sequence

Fig. 13- 1 2 3 1 2 4 8 PCR cycles DNA copies 4 16 etc. etc. DNA fragment to be amplified (90°C) 122°F (50°C) 158°F (70°C) DNA polymerase new DNA rimers strands ating ates trands Cooling allows primers and DNA polymerase to bind New DNA strands are synthesized 2 3 le (b) Each PCR cycle doubles the number of copies of the DNA Each PCR cycle doubles the number of copies of the DNA

Author Animation: Polymerase Chain Reaction

(PCR)

 (^) Differences in short tandem repeats can identify individuals by their DNA

• (^) Forensic scientists have found that small, repeating segments of DNA, called short tandem repeats (STRs) can be used with astonishing accuracy to identify people - (^) Each STR is short (consisting of 2 to 5 nucleotides), repeated (as many as 5 to 50 times), and arranged in tandem (back-to-back)
• (^) STRs are repeated sequences of DNA within the chromosomes that do not code for proteins (INTRON regions)

 (^) Differences in short tandem repeats can identify individuals by their DNA (continued)

• (^) STRs vary greatly between different human individuals, like genetic fingerprints
• (^) Different people may have different alleles of the STRs (i.e. differing # of repeats)
• (^) Law Enforcement Agencies have established a standard set of 13 STRs, each four nucleotides long, to identify individuals by DNA samples - (^) There may be as few as 5 to as many as 38 repeats of these STRs
• (^) Commonly referred to as “DNA fingerprinting”

Short-Tandem Repeats Are Common in Noncoding

Regions of DNA

Fig. 13-4 Eight side-by-side (tandem) repeats of the same four-nucleotide sequence

 Gel electrophoresis separates DNA segments

• (^) Mixtures of DNA fragments can be separated on the basis of size
• (^) Gel electrophoresis is a technique used to spread out DNA fragments of varying lengths in a mixture
• (^) DNA is placed at one end of a gel
• (^) A current is applied to the gel
• (^) DNA molecules are negatively charged and move toward positive end of gel
• (^) Smaller molecules move faster than larger ones

 Gel electrophoresis separates DNA segments

(continued)

• (^) There are four steps in gel electrophoresis
  1. DNA mixtures are placed into wells at one end of a slab of agarose gel, and an electric current is introduced to the gel
  2. Negatively charged DNA moves toward the positive electrode
     • (^) smaller fragments moving through the gel meshwork more quickly than larger ones
     • (^) mixture is thus separated into bands of DNA along the gel, according to fragment lengths