Advanced techniques in molecular biology

Question 1

Steps of molecular phylogenetics

Answer 1

1. alignment: building the data model and extracting a dataset
2. determining the substitution model
3. consider sequence variation
4. tree building
5. tree evaluation

Question 2

Methods to manipulate gene expression

Answer 2

• RNAi
• over expression

Question 3

Methods used for local studies

Answer 3

• IHC
• quantum dots

Question 4

Methods used for functional studies

Answer 4

• co-immunoprecipitation
• proximity ligation assay (PLA)
• FRET
• FRAP (fluorescence recovery after photobleaching)

Question 5

non-coding RNA (ncRNA)-associated gene silencing can be done by

Answer 5

• chromatin remodelling
• transcriptional regulation
• post-transcriptional regulation

Question 6

Microscopy methods

Answer 6

• fluorescence microscopy
• confocal m.
• electron m.
• atomic force m.

Question 7

Name some basic molecular biology techniques

Answer 7

• DNA cloning
• PCR
• gel electrophoresis
• molecular hybridisation
• mutagenesis
• DNA and protein arrays

Question 8

Steps of protein modeling / molecular modeling

Answer 8

1. identify related proteins with well-known 3D structures -
2. align the related 3D structures with the target sequence and determine those structures that will be used as a template -
3. contruct a model for the target sequence based on its alignment with the template structure(s) -
4. evaluate the model against a variety of criteria to determine if it is satisfactory

Question 9

What detection techniques can be used in stainings?

Answer 9

• Fluorophore or fluorochrome
• DAB: 3,3’-diaminobenzidine tetrahydrochlorind
• AEC: aminoethly carbazole
• AP: alkaline phosphatase

Question 10

Name some advantages of quantum dots over conventional fluorophores.

Answer 10

• They are more stable
• they have narrow spectra
• they shine brighter
• they last longer
• are often less toxic
• specific organic molecules can be attached to them

Question 11

It is proposed that histone modifications affect chromosome functions. What are the two mechanisms that are suggested to do this?

Answer 11

1. modifications may alter the electrostatic charge of the histone resulting in a structural change in histones or their binding to DNA.
2. the modifications are binding sites for protein recognition modules that recognise acetylated or methylated amino acids