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BIOC0005 > Nucleic Acids > Flashcards

Nucleic Acids Flashcards

1
Q

What is a nucleoside?

A

A purine or pyrimidine base, bonded to a sugar molecule by a glycosidic bond.

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2
Q

What is a purine, and name the purine bases.

A

A double ringed base.

Adenine and Guanine.

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3
Q

What is a pyrimidine and name the pyrimidine bases.

A

A single ringed base.

Cytosine, Thymine and Uracil.

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4
Q

Compare the labelling of the carbon atoms in the base vs the sugar molecule.

A

Base- carbon atoms are numbered from 1-.

Sugar- carbon atoms are numbered with prime.

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5
Q

What is a nucleotide?

A

Compounds consisting of a nucleoside linked to a phosphate group (s).
These form the basic structural units of nucleic acids DNA and RNA.

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6
Q

What are nucleic acids?

A

Polymers of nucleotides.

Molecules that allow organisms to transfer genetic information between generations.

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7
Q

Compare DNA to RNA.

A

DNA-

  • Deoxyribose sugars.
  • No uracil base.
  • Double stranded.
  • Storage form of the genetic information.
  • Heteropolymer of 4 different nucleotides (ATCG).
RNA- 
-Oxyribose sugars. 
-Uracil bases instead of Thymine.  
-The working copy of the gene.
-Single stranded. 
Heteropolymer of 4 different nucleotides (AUCG).
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8
Q

What is a glycosidic bond?

A

It is the bond formed between the sugar molecule and the base. Water molecule is released.

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9
Q

Between which atoms is the glycosidic bond formed?

A

1’ of the carbon of the sugar molecule, and the N-1 (pyrimidines), or N-9 (purines).

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10
Q

What enzyme is involved in nucleotide polymerisation?

A

DNA polymerase.

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11
Q

Outline the process of nucleotide polymerisation.

A

DNA polymerase, forms hydrogen bonds and covalent bonds in the 5’ to 3’ direction through the polymerisation of the monomer dNTPs.

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12
Q

Where are the hydrogen bonds formed in DNA?

A

Between the complementary bases of the two strands.

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13
Q

How is the geometry of the DNA helix maintained?

A

Through the hydrogen bonds.
They are formed between a purine and pyrimidine.
GC- 3x H-bonds.
AT- 2x H- bonds.

Know structure of bonding!

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14
Q

Where are covalent bonds formed in the DNA molecule?

A

Between the sugar and phosphate of adjacent bases, to form the phosphodiester backbone… 5’ phosphate, and 3’ OH.

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15
Q

How was the 3D structure of DNA discovered?

A

By x-ray diffraction photographs in 1953 by Watson and Crick.

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16
Q

What is Chargaff’s rule?

A

The CG, AT base pairing rules… to maximise the number of effective hydrogen bonds that can form between bases, allowing formation of the most stable conformation.
Pyrimidine always paired with a purine to keep the same distance between the two strands so that the bases can bond, maintaining the geometry.

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17
Q

What is the importance of minor and major groves?

A

I.e. where the backbone is closer/ further apart.
Major grove… more exposed… allows certain proteins to bind to DNA to alter its structure or to regulate transcription or replication.

18
Q

What are the three DNA helix geometries?

A

A-DNA.
B-DNA.
Z-DNA.

19
Q

What is the most predominant type of structural conformation of DNA in the cells?

A

B-DNA, occurring under normal physiological conditions.

20
Q

Outline how DNA is packaged.

A
  • A double stranded DNA helix is complexed with histones to form nucleosomes.
  • Each nucleosome consists of 8 histone proteins around which the DNA wraps 1.65 times.
  • A chromatosome consists of a nucleosome plus the H1 histone.
  • The nucleosome folds up to produce a 30n m fibre, forming loops around 300nm in length, which are compressed and folded to produce a 250nm-wide fibre.
  • THe tight coiling of the 250nm fibre produces the chromatid of a chromosome.
21
Q

Compare DNA packaging in eukaryotes vs prokaryotes.

A

Eukaryotes-

  • One long, linear dsDNA molecule.
  • Bound to a complex mixture of proteins to form chromatin.
  • Closed, circular ds DNA molecules in mitochondria and in chloroplast.

Prokaryotes-

  • Single, circular, dsDNA molecule.
  • Can take the supercoiled, circular, or nicked form, although can also undergo positive and negative supercoiling.
  • Generally prokaryotic chromosome is associated with non-histone that help bacteria to compact the DNA to form a nucleoid.
  • Extrachromosomal DNA plasmid.
22
Q

What is a nuclease?

A

Enzyme that can cleave the phosphodiester bond between the monomers of nucleic acids, at specific site.

23
Q

What is an exonuclease?

A

Works by cleaving nucleotides one at a time from the end of the polynucleotide.

24
Q

What is an endonuclease?

A

Enzyme that cleave the phosphodiester bond in the middle of the polypeptide chain.

25
Q

What is a restriction endonuclease and name 3 different ones.

A

An enzyme that cleaves both strands of dsDNA at a specific sequence… staggered cuts.
Palindromic recognition site.
EcoR1, Pst1, Hpa1.

26
Q

What is a palindromic sequence?

A

A sequence that reads the same forwards as backwards.

27
Q

List some characteristics of the EcoR1 enzyme.

A
  • Recognises the palindromic sequence GAATTC.

- Forms sticky ends with a 5’ overhang.

28
Q

List some characteristics of the Pst1 enzyme.

A

Recognises specific palindromic sequence CTGCAG.

- Forms 3’ overhangs.

29
Q

List characteristics of the Hpa1 enzyme.

A
  • REcognises specific palindromic sequence GTTAAC.

- Cuts betten AT in the middle, forming a blunt end cut.

30
Q

What is alkaline phosphatase?

A

An enzyme with the physiological role of dephosphorylating compounds.

31
Q

What is alkaline phosphatase used for in the lab?

A

To remove phosphate monoesters, to prevent self-ligation… undesirable in plasmid DNA cloning.

32
Q

What are DNA ligases?

A

Enzymes that join DNA, and will catalyse the formation of a phosphodiester link.
Sticky end ligation, bringing complementary strands together.

33
Q

Why is blunt end ligation less efficient ?

A

As you are just bringing two pieces of DNA together i.e. can’t match up as efficiently as DNA with sticky ends, so need more DNA to be put in the tube, but can be useful when the sticky ends can’t be used.

34
Q

How can DNA strands separate?

A

DNA denaturation, enzymes, in the lab through heat and high pH.

35
Q

What is the Tm of DNA?

A

The melting temperature i.e. the midpoint… temperature at which the strands separate, i.e. can overcome the hydrogen bonds.

36
Q

What factor affects the DNA’s Tm?

A

The sequence composition of DNA, i.e. CG has three hydrogen bonds so stronger, need more heat to separate, so higher Tm.

37
Q

What is hyperchromicity?

A

A material’s increasing ability to absorb light.
I.e. when DNA goes from an ordered to a disordered structure, the absorbance (A) at 260nm increases. I.e. when DNA is heated… denatured.
The increase of absorbance (optical density) of a material.

38
Q

What is hybridisation?

A

Complementary base pairings between DNA fragments, can form in different combinations.

39
Q

What is the Southern Blot?

A

Once DNA digestion is run on agarose gel and visualised under UV light, transferred DNA from gel onto membrane, then add ssDNA gene probe that is complementary to the gene you are looking for… cloned fragment. Binds to the complementary strand, so can develop autoradiography… black line where probe has bound to complementary DNA.

40
Q

What is DNA topology?

A

The coiling of a molecule of DNA.

41
Q

How are different plasmid topoisomers separated by gel electrophoresis?

A

In order of distance travelled (low- high)… Relaxed/ open circle, linear, supercoiled.

BIOC0005 (9 decks)

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