Unit 13

Biotechnology

Biotechnology

• The use of cells and biological molecules to solve problems or make useful products

• Involves manipulation of DNA and RNA

DNA

• Building block of DNA is a nucleotide– Deoxyribose, phosphate group, and a base

• Bases are adenine, thymine, guanine and cytosine.

• Two strands of DNA molecules are held together by hydrogen bonds which form between the bases– A-T– G-C

Figure 11.7 Base Pairing in DNA Is Complementary

Phosphodiester bonds link the sugar and phosphate

Hydrogen bonds hold strands together by complementary base pairing

DNA replication

• DNA helix unwinds

• Each strand serves as a template for a new strand

• How would you replicate this strand of DNA?– TTT GCC AGA

• Tutorial 11.2 (DNA replication)

Transcription

• Process of producing a messenger RNA copy using the DNA template.

• mRNA takes the message from the nucleus to the cytoplasm

• In RNA, there is no thymine (T)• In RNA, sugar is ribose, not deoxyribose• Base-pairing in RNA

– A-U– C-G

• How would you transcribe this DNA?– ATC GGG TTA

• Tutorial 12.1

Translation

• Messenger RNA is “translated” into protein at the ribosome

• The 3-letter triplet codon on mRNA directs transfer RNA to bring the correct amino acid to the growing chain

• Fig 13.7 shows the genetic code• What amino acids would be added to the

chain based on these codons?• AUG CUU UGU GCG

Figure 12.5 The Universal Genetic Code

• Steps in translation are initiation, elongation and termination

• View animation of translation and protein synthesis

• http://www.whfreeman.com/thelifewirebridge2/ tutorial 12.3

Exercise 13.1

• DNA puzzle

• Simulate DNA replication

• Simulate transcription– Be sure to use ribose sugar instead of

deoxyribose– Different colors for ribose and deoxyribose

• Simulate translation

Exercise 13.2

• Isolation of DNA from onion or banana cells

• Filtrates will be prepared and students will add the ice-cold ethanol to precipitate the DNA.

• Observe DNA as a stringy, cottony mass.

Exercise 13.3• Restriction enzymes

– Enzymes that recognize a particular DNA sequence and cut there

• Example EcoRI– Restriction site is

G A A T T C

C T T A A G cuts between the G and the A

• The enzyme EcoRI cuts DNA with the following paired sequence:– 5 ... GAATTC ... 3– 3... CTTAAG ... 5

• Notice that the sequence is palindromic: It reads the same in the 5-to-3 direction on both strands.

Figure 16.4 Cutting and Splicing DNA

Fragments from a human source can be joined to another source, such as a bacterium

• Some restriction enzymes cut DNA strands and leave staggered ends of single-stranded DNA, or “sticky” ends, that attract complementary sequences.

• If two different DNAs are cut so each has sticky ends, fragments with complementary sticky ends can be recombined and sealed with the enzyme DNA ligase.

• These simple techniques, which give scientists the power to manipulate genetic material, have revolutionized biological science in the past 30 years.

• Hundreds of restriction enzymes have been purified from various organisms, and these enzymes serve as “knives” for genetic surgery.

• The fragments of DNA can be separated using gel electrophoresis. DNA is negatively charged

• When DNA is placed in a semisolid gel and an electric field is applied, the DNA molecules migrate toward the positive pole.

• Smaller molecules can migrate more quickly through the porous gel than larger ones.

• After a fixed time, the separated molecules can then be stained with a fluorescent dye and examined under ultraviolet light.

Figure 16.2 Separating Fragments of DNA by Gel Electrophoresis (Part 1)

Figure 16.2 Separating Fragments of DNA by Gel Electrophoresis (Part 2)

Figure 16.2 Separating Fragments of DNA by Gel Electrophoresis (Part 3)

Kitchen electrophoresis

• Prepared samples will be loaded onto an agarose gel by instructor and electrophoresed.

• You will use the crime scene scenario to interpret the gel. Which sample matches with the one taken from the crime scene?– Compare pattern of bands. If pattern of DNA bands from the

crime scene matches the pattern of one of the suspects, this provides evidence that the suspect is guilty.

• Also view gels showing results of paternity testing.– View the patterns for the mother, child and the potential fathers

to determine which is likely to be the father of the child.

Agar plate inoculation for next week

• Prepare agar plates by swabbing with environmental samples.

• These plates will be viewed next week.• Draw lines on bottom of plate (not the lid)

to divide plate into 3 sectors.• Swab 2 sectors with different

environmental samples and label. • Leave one sector as a control.• Tape down the lid and leave on cart.

Check-out procedures

• Demonstrate transcription and translation using puzzle pieces

• Explain crime scene interpretation