Synthetic biology

Question 1

Synthetic biology has a bottom-up approach. What does this mean?

Answer 1

It tries to assess whether complex systems can be made from parts, and whether they can be redesigned.

Question 2

Give examples of synthetic biology.

Answer 2

1. Engineered enzymes
2. Gene circuits
3. Custom genomes
4. Unnatural substrates

Question 3

In synthetic biology systems are broken down into parts. What might serve as the ‘chassis’?

Answer 3

The organism of interest, e.g. bacteriophages, bacteria, yeast etc.

Question 4

Define a BioBrick.

Answer 4

DNA sequences that conform to a restriction-enzyme assembly standard.

Question 5

What are BioBricks used for?

Answer 5

To design and assemble synthetic biological circuits. These are then incorporated into living cells to build new systems.

Question 6

How are new parts introduced, i.e. how are BioBricks joined?

Answer 6

Wit restriction endonucleases. The original restriction site changes with each addition of a new brick.

Question 7

What do the assembled parts generate?

Answer 7

A device, which itself is a composite part.

Question 8

Where are new parts added?

Answer 8

At the ends of the starting device/part.

basically can only be added in order

Question 9

Parts can be tested to evaluate robustness and reproducibility before they are integrated into living cells. True or false?

Answer 9

True.

Question 10

Define a logic gate.

Answer 10

It performs a logical function on one or more inputs. This produces a single output that is a logical function of the inputs.

Question 11

Logic gate equivalents are used in synthetic biology, what are the components?

Answer 11

Paths for biological molecules to travel down (wires equiv.) and places where they can pass or be blocked.

Question 12

Complex circuits are just as robust as simple circuits. True or false?

Answer 12

False: complex circuits are far less robust than simple circuits.

Question 13

Can devices be integrated between cells?

Answer 13

Yes.

Question 14

Give 2 advantages of BioBricks.

Answer 14

1. Standardised parts can be created and shared

2. Much of the design process can be automated, causing fewer errors

Question 15

Give 3 disadvantages of BioBricks.

Answer 15

1. When all parts are viewed as independent modules then context is ignored, meaning there is less robustness
2. Scars are created at junctions when different BioBricks are ligated together.
3. Serial assembly is slow even when automated.

Question 16

Define a scar.

Answer 16

When BioBricks with different RE sites are joined, both palindromic sequences are destroyed. Thus the 2 bricks can never be cut apart. This mixing of RE sites forms a scar.

Question 17

When can scars be a problem?

Answer 17

If the device is being used to create a single polypeptide. The scar is a bit of mis-sense DNA.

Question 18

Define a Type IIS RE.

Answer 18

Cut outside of their asymmetric, non-palindromic recognition sites.

Question 19

How can Type IIS REs be used in scarless cloning?

Answer 19

They do not create scars when 2 BioBricks are joined as the recognition sites are non-palindromic. Plus it does not require the introduction of new cutting sites.

Question 20

Define Gibson assembly.

Answer 20

Allows the joining of multiple DNA fragments in a single isothermal (temperature remains constant) reaction.

Question 21

What 3 kinds of enzymes, other than REs, does Gibson assembly require? Briefly state why they are needed.

Answer 21

1. Exonucleases: unmask compatible ends
2. Polymerases: fills in gaps between free 3’ and 5’ ends
3. Ligases: assembles the construct into a single molecule

Question 22

Gibson assembly produces scars. True or false?

Answer 22

False.

Question 23

Gibson assembly allows multiple fragments to be joined at once. True or false?

Answer 23

True.

Question 24

Gibson assembly is used to assemble small constructs. True or false?

Answer 24

False: it is used for larger constructs.

Question 25

What is the term ‘Synthia’ used to describe?

Answer 25

Synthetic life.

Question 26

Multiple fragments can be assembled in vivo. How?

Answer 26

Using yeast recombination machinery.

Question 27

Biological systems result from evolutionary processes. Why are components in their current arrangement? There are 2 possibilities.

Answer 27

1. Intrinsic limitation
2. Frozen accident

Synthetic biology seeks to find out and thus improve upon systems.

Question 28

Information is stored in DNA but only accessible via RNA. True or false?

Answer 28

True.

Question 29

All 3 chemical groups of a nucleotide - the sugar, the nitrogenous base and the phosphate - contribute to its structure and function. True or false?

Answer 29

True.

Question 30

What is XNA?

Answer 30

‘Xenobiotic nucleic acids’: synthetic nucleotides whereby one or more of the 3 distinct chemical groups have been modified.

Question 31

XNA have variable properties and compatibility with biological systems. True or false?

Answer 31

True.

Question 32

Define an episome.

Answer 32

Non-integrated, extra-chromosomal DNA.

Question 33

If XNA is used as the genetic material, what may we be able to create in the future?

Answer 33

XNA episomes.

Question 34

Why might an XNA episome be useful? Give 2 examples.

Answer 34

1. Can store extra information: if we establish a coding/decoding mechanism with synthetic enzymes whereby XNA and DNA are interchangeable.
2. Can be maintained stably as is not part of the chromosomes.

Question 35

On what condition would and XNA episome need to work?

Answer 35

Orthogonality: XNA would need to be inaccessible to current biological systems (basically so it doesn’t fuck your cells up), i.e. they can both occur without interfering with each other.

Question 36

Define Xenobiology.

Answer 36

Unnatural biological systems.

Question 37

What are the risk of GMOs resulting for xenobiology? Give 2 examples.

Answer 37

1. Ecological: GMOs may interfere with other animals and their niches.
2. Informational: the risk that part of the GMed genome could spread to natural organisms, which would disrupt ecosystems.

Question 38

Reports of controlled GMO release have so far conferred what?

Answer 38

GMOs released for bioremediation have not established themselves in the tested niches and pose no threat to the environment.

Question 39

XNA can be viewed as a ‘dead man’s switch’. Why?

Answer 39

Because it requires synthetic prescursors and xenobiology is purposely isolated from natural biology, it does not require an organisms to be alive to be activated, e.g. does not require precursors to be synthesised etc.

Question 40

Containment failure of XNA relies on the ‘shortest evolution distance’. What does this mean?

Answer 40

The smallest change could cause it to be uncontained: for example a single mutation is archaea with XNA caused the production of XNA reverse transcriptase, causing RNA to be produced from XNA.