Genetics Ramaswami lectures 3 & 4

what is the name of the short DNA sequences which can be chemically synthesised?

oligonucleoties

are oligos usually single or double stranded?

single

how long are oligos usually?

21 to 29 bases

outline the 3 major steps of PCR (denaturation, annealing and extension)

1. denaturation by increasing the temp to 95C to separate the two DNA strands

2. annealing of primers by reducing the temp to 50-60C

3. extension of a new complementary dna sequence by dNTPs and Taq pol

amplification of the target DNA proceeds … in PCR

exponentially

what technique is used for rapid diagnosis of viruses which use reverse transcriptase?

rRT PCR

rRT PCR amplifies what?

cDNA produced by reverse transcriptase from viral mRNA

which enzyme is used to convert the viral RNA into cDNA which is then amplified with PCR and used during fluorescence based product detection to identify infections of SARS CoV 2?

reverse transcriptase

what genome does SARS CoV 2 (virus which causes covid 19) have?

+ssRNA

in real time fluorescence detection:

1. a fluorescent … or … binds only to the amplified target dsDNA

2. the PCR machine measures … after each cycle, so you see the amplification curve in real time

1. dye or probe

2. fluorescence

in quantification of real time fluorescence detection: the lower the number of cycles needed to reach a certain fluorescence threshold, the higher the levels of … in the sample

viral RNA

PCR is used in genotyping of alleles which show what kind of polymorphism?

length polymorphism

what does VNTRs stand for

variable number of tandem repeats

region of DNA where a sequence is repeated multiple times

tandem repeat

are tandem repeats found in the coding or noncoding DNA?

noncoding

are short tandem repeats (STR) classified as micro or minisatellites?

microsatellites

are variable number tandem repeats (VNTR) classified as micro or minisatellites?

minisatellites

any tandem repeat where copy number varies (VNTR/STR)

VNTR

a tandem repeat where te sequence repeated is short (ie 2-6 bases) - is a subtype of VNTR

STR

for PCR we have to design … and … primers for the gene of interest

forward and reverse

a major application of PCR

genotyping of alleles based on the allele specific length differences

what technique is used to separate different amplified target alleles based on their length

gel electrophoresis

what type of genotyping uses fluorescently labelled primers designed to amplify alleles of different sizes (which can also be separated on the same gel)?

multiplex

what type of genotyping produces this gel electrophoresis?

multiplex

multiplex genotyping allows alleles at many different … to be investigated at the same time

loci

areas of major applicationf of multiplex genotyping

1. … science

2. … testing

3. … genetics

4. … genetics

1. forensic science

2. paternity testing

3. population genetics

4. evolutionary genetics

what kind of nucleotides can terminate chain synthesis?

dideoxynucleoties

which is faster next generation sequencing or chain termination sequencing?

next generation

is it sanger or next generation sequencing where billions of strands of DNA are sequenced at once?

next generation

is it sanger or next generation sequencing where only one strand of DNA is sequenced at once?

sanger

the DNA analysis method where a long DNA strand is randomly broken into many small, overlapping fragments, each sequenced individually, and then reassembled by computers using the overlaps to reconstruct the original, full sequence

shotgun sequencing

the DNA/RNA sequencing technology that rapidly reads millions of genetic fragments in parallel

next generation sequencing

what are genomics?

the study of the entire set of DNA of an organism and its genes, exploring how they function, interact and influence traits

human genomics are useful to identify … which cause specific diseases, disease …, drug … and traits

answer with a list

1. mutations which cause specific diseases

2. disease resistance

3. drug sensitivity

4. traits

suggestive evidence for RNA being the intermediate between DNA and proteins

1. DNA is located in the nucleus

2. RNA is synthesised in the nucleus

3. RNA migrates to the cytoplasm where proteins are synthesised

4. in viral infection RNA synthesis precedes new viral particle formation

at which position does deoxyribose lack OH at?

2

what does UMP stand for?

uridine monophosphate

what does AMP stand for?

adenine monophosphate

what does CMP stand for?

cytidine monophosphate

what does GMP stand for?

guanosine monophosphate

name all four nucleotides in RNA

adenine, cytidine, guanosine and uridine monophosphate

which type of RNA represents <10% of RNA in a cell?

mRNA

does mRNA have a long or short half life?

short

what is the most abundant RNA in cells?

rRNA

which type of RNA represents >90% of RNA whithin a cell?

rRNA

rRNA represents a … and … component of ribosomes

structural and functional

which type of RNA is the smallest?

tRNA

which type of RNA is less than 100 bases long?

tRNA

do prokaryotes have one or multiple RNA polymerases?

one

what are the three RNA polymerases in eukaryotes?

RNAP I, RNAP II and RNAP III

which RNA does RNAP I make?

rRNA

which RNA does RNAP II make?

mRNA

which RNA does RNAP III make?

tRNA

does the template or nontemplate strand get transcribed?

template

is the coding strand the same as the template or nontemplate strand?

nontemplate

is it the template or nontemplate strand which has the same nucleotide sequence as the mRNA being produced except T → U

nontemplate

is it the template or nontemplate strand which is the antisense?

template

template = …

nontemplate (coding) = …

(sense/antisense)

template = antisense

nontemplate = sense

the DNA sequences which guide RNAP to the beginning of the gene to start transcription

promoters

the DNA sequences that specify the termination of transcription and release RNAP from the DNA

terminators

3 steps of transcription (just 3 words)

1. initiation

2. elongation

3. termination

what number denotes the transcription initiation site?

+1

is there a 0 position relative to the trascription initiation site of +1?

no

is the upstream sequence denoted by +/-?

+

is the downstream sequence denoted by +/-?

-

what is another name for the TATA box in prokaryotes?

pribnow box

how many bases upstream is the TATA/Pribnow box in prokaryotes?

-10bp

what is the other upstream box in prokaryotes?

-35bp box

prokaryotic promoters contain 2 highly conserved regions.

copy this cus its important

prokaryotic promoters contain 2 highly conserved regions

how long are prokaryotic promoters typically?

40bp