dna and rna synthesis Flashcards
What is the role of genetics in periodontal disease?
The interleukin-1B gene variation causes overproduction of interleukin-1
In bacterial plaque- high inflamm response leading to destruction of soft tissue and bone
What is DNA fingerprinting?
Relies on variable no of tandem repeats (30-40% of non coding region) Myoglobin gene- segments in introns w no apparent function- vary in size and composition between individuals Initially called mini-satellites as surround the exon Repeating nucleotides (15-200), no of repeats and loci inherited Large variation in no of regions between people
What is VNTR?
Variable number of tandem repeats
Found at single/many locus/loci
How does PCR work ?
Polymerase chain reaction
Measures transcription of gene
Uses DNA
Reverse transcription- use RNA to make cDNA and use PCR to make many copies of specific piece of DNA
Can detect w lots of DNA
How was PCR testing established?
1955- first DNA polymerase
1985 Mullis- developed polymerase reaction
Thermostable polymerase, Taq, enzyme from Thermus aquaticus (bacteria living in hot water 72C)
Components needed for PCR? ~template ~primers ~A,C,G,T ~Mg and Taq
How does PCR work?
DNA separated by heating 94C Add primers to stick New DNA is made via Taq and by adding magnesium and A,G,C,Ts Heat to 72C Usually 35 cycles and 68B copies
What are problems w amplifying sequences?
Poor precision Lower sensitivity Short dynamic range <2 logs Low resolution Non automated Size based discrimination only Not quantitative Ethidium bromide staining not quantitative
What fluorescence is used to count genes?
SYBR green- binds to any double stranded DNA
TAQ polymerase- requires specific probe to bind specific sequence
What are the advantages of real-time PCR?
Collects data in exponential growth phase
Increase in reporter fluorescence directly proportional to no of amplicons generated
Increased dynamic range of detection
No post PCR processing
Detection down to 2-fold change
What is in situ hybridisation?
Localisation technique
Detects gene product in tissues
What is genomics?
DNA/RNA sequencing and bioinformatics
Used to sequence, assemble and analyse the function and structure of genome
What is Oxford nanopore?
1-2 days to sequence DNA/RNA
Reads 400 based per second
What are transcriptomics?
Genome wide RNA transcript expression levels
Next generation sequencing m
What are proteomics?
Large scale study of proteins, esp structures and functions inc post translational mods
What are metabolomics?
Characterisation of small molecule metabolites on bio systems
