TOPIC E: DEVELOPMENT Flashcards
what are descriptive studies based on?
- Expression profiles of mRNA
- Non coding RNAs and proteins
- Temporal (when) and spatial (where)
- fate mapping of cells (lineage tracing)
- comparative studies of organisms
What are manipulative studies and 3 examples?
ALTERING SYSTEM AND OBSERVING EFFECTS
- Gene knockout or transgenic overexpression
- Adding drug or tetragon and observing effects
- Translating cells from ONE area to ANOTHER (cell commitment and specification)
What kind of microscopy is a light microscoope used for?
- Brightfiled microscopy
What kind of microscopy is a confocal microscope used for?
- Fluorescence microscopy
What kind of microscopy is an electron microscope used for?
- TEM (Transmission electron microscopy–> can “see” inside cell)
Does temporal gene expression analysis in tissue examine WHEN a gene is expressed?
- YES “when”= time
What does DNA analyis include?
- PCR (promoter and enhancer studies–> plasmids)
- gene cloning (epigenetics–> methylation of DNA, histone modification)
- Southern blotting– >gene knockout or overexpression
What does RNA analysis include?
- gene expression analysis
- RT-PCR
- RNA -seq
- Knockdown of mRNAs using RNA intereference (RNAi)
What does protein analysis include?
- Western blotting and immunostaining of tissues
- Protein structure –> function, phosphorlyation, sumoylation (to do with ubiquitin) , acetylation, glyocsylation
- Protein-protein interactions by co-immunoprecipiation
WHat does bioinformatics include?
- using computer software to understand biologcial data
e. g. constructing whole genomes, comparing data genes or sets
What is real time PCR?
- Quanititative Reverse Transcription -PCR (q RT-PCR)
- converts RNA—> cDNA (via reverse trancription)
- Fluorescent ATCG labells added to mix
What do a lower number of PCR cycling to reach detection indicate?
- More starting RNA
What do a higher number of cycles of PCR cycling to reach detection indciate?
- Less cDNA present therefore less RNA
What does in situ hybridisation look at and what colour stain?
- Study of prenatal development
- temporal and spatial expression of mRNAs in MODEL embryos (fly, fish, chicken, mouse)
- Purple stain for cells with the RNA of interest
What is imunofluorescence to do with?
WHEN and WHERE a protein is epxressed in a tissue (antibodies against protein of interest)
What does Western Blotting show in terms of the proteins?
- The SIZE of the protein and the relative amount present in a tissue
Does the primary antibody for Western blotting or Immunofluorescne have to be made in the same animal or separate animals?
- Separate animals for different cell strucutres
Does a second degree antibody have to be made in separate animals to target different primary antibodies?
- NO
- Can be made in SAME animal
What does a transgenic mice model involve?
- Carrying foreign gene and we observe the phenotype
What are the 2 methods for creating trasngenic mice?
- Pronuclear injection
2. Transforming ES cells (embryonic stem cells)
What does modern developmental bio involve?
- Embryology
- Cytology (cells ) + Histology (tissues)
- genetics
- understands the genetic and cell mechanisms that allow 1 cell–> multicomplex cell
What issues does Dev bio help with?
- Cancers, cell therapy for human disease (stem cells) , developmental disorders `
What is pattern formation?
- Once cell–> many cells–> organised cells–> tissues–> organs
Which two processes does pattern formation require?
- Differential gene expression
2. Signalling between cells