DNA genetics Flashcards
How goes the information on DNA determine protein structure?
dsDNA has a template strand (3’-5’), and an RNA-like strand (5’-3’). The template strand has a complimentary sequence to the RNA like strand of the mRNA-the sense strand or + strand. RNA polymerase goes from 3’ to 5’ on the template strand producing mRNA going from 5’ to 3’ identical to the RNA-like strand. The 5’ start codon of mRNA when translated becomes the N-terminal end of the protein transcript, 3’ end the C terminal.
What are long non-coding RNA (IncRNA)?
Long non-coding RNA (>200 bp) has been found to modify gene expression that can be inhibited by small interfering RNA. Protein coding RNA accounts for only 1/5 of transcription. IncRNA occurs in osteoarthritis and encourages metalloproteinase production and inhibition results in increased aggregation.
Why are some loss of function mutations dominant?
A dominant allele usually codes for a functional protein, and other alleles are recessive as the altered protein is less functional. Loss of function occurs in a diploid organism if the gene product of the wild type allele is insufficient for normal function, resulting in an abnormal phenotype due to the non functional recessive allele. The mutation is due to haploinsufficiency eg hypercholesterolemia. If the mutation antagonizes the normal product the genotype is called a dominant negative mutation eg Huntington’s chorea.
How does miRNA interfere with DNA transcription?
The miRNA-inducing silencing complex (RISC) is generated by Drosha binding of primary transcript and cropping, exporting to cytoplasm where Dicer crops dsRNA then degrades one strand producing miRISC, which pairs with messenger RNA causing cleavage and mRNA destruction or translation inhibition. if only partly complementary.
What role might endogenous retroviruses play in gene expression?
Endogenous retroviruses present in eukaryotic genomes may function as promoters and rarely expressing their own genes. Salivary amylase owes it’s expression in salivary tissue to an endogenous retroviral promoter. Placental function depends upon similar retroviral effects. ERV account for about 9% of mammalian DNA.
How many types of transposable elements are there in the human genome ? How common are they?
There are 2 categories transposable elements making up 45-48% of human genome:-
1) Retrotransposons (RNA intermediate)
a) SINEs (short interspersed nuclear elements) 1-5Kb (U3-R-U5 structure at both ends of retroviral sequences), have poly A tail, 2 million per genome, ~11% human genome.
b) LINEs (long interspersed nuclear elements), contain ORF1, and pol genes, poly A tail, 6-8 kb long, 1 million per genome. 21%.
c) HERVs (Human Endogenous Retrovirus), gag, pol and env genes plus LTR’s at both ends. 1-11 kb long, 600,000/genome, 8%.
2) DNA transposons, have no RNA phase, contain 1 transposase gene flanked by inverted repeats, 350 base pairs long, 400,000/genome, 3%. Some LINES are autonomous (100/genome), can move by themselves. All SINES are non autonomous and require expression of LINE genes. Almost all LINE and SINE insertion points are the same in all humans with 8000 mobilized during human history. Alu elements are primate retrotransposons, 300 bp, (SINE), make up 11% human genome, of which 7k are unique to humans.
Composite transposons formed by fusion of Alu and SINE-R are the most recent and found in gorillas, orangutans and humans. Helitrons make up 0-3% of mammalian genomes which utilize a single strand DNA intermediate and transpose via rolling stone replication.
What is the difference between ordinary PCR and ms-pcr?
Ordinary PCR produces amplicons - more than one copy of a genetic fragment of a target sequence. ms-pcr refers to methylation-specific polymerase chain reaction which detects methylation sites. Treatment with bisulphite changes non methylated cytosine to uracil, methylated cytosine is resistant.
What is a CpG motif?
CpG motifs refers to a DNA sequence where cytosine is 5 prime to quanine. The cytosine can be methylated to methylcytosine in mammals by DNA methyltransferases which turns the gene off.
What is the difference between a promoter and enhancer?
A promoter is a region of DNA that initiates transcription of a particular gene. They are located near the start site on the same strand and upstream of the 5’ region of the sense strand and can be 100 to 1000 base pairs long. TATA box binding proteins (TBP) and TBP associated factors (TAF’s) bind the promotor sequence then attract pol II to start transcription. An enhancer is a DNA sequence that is trans activated by transcription factors, on the cis-acting enhancer sequence, located up to 1 million base pairs (1Mbp) away upstream or downstream from the start site. TF binding attracts mediator proteins that complex with the pol II complex by looping the DNA strand. Transcription increases by changing the polymerase and attracting co-activators which remove nucleosomes. Insulator sequences work by fixing loop formation that prevents enhancer looping.
How does the REST/NSRF repressor protein work?
REST/NSRF (RE1-silencing transcription factor/neuron-restrictive silencer factor) is a repressor protein, 1,097 aa with 9 zinc finger motifs, that associates with mSin3 (interaction domain for TF suppression) and CoREST (nerve differentiating factor) to silence many transcription factors by deacetylating and demethylating specific sites on histones. Active in neurons, and inactive in neoplasia-small cell lung CA.
Where does the internal ribosomal entry site (IRES) usually insert?
An internal ribosome entry site (IRES) is a nucleotide sequence that permits ribosomes to begin at that point on mRNA. IRES are often located on the 5’ UTR of RNA viruses, and in mRNA of animal genes.
What is the function of terminal deoxynucleotidyl transferase?
Terminal deoxynucleotidyl transferase adds N-nucleotides to the V, D, and J exons during antibody gene recombination thus enabling junctional diversity. The DNTT gene is active in immature T and B cells and multipotent hematopoietic stem cells. KO mice show a 10 fold reduction in T-cell receptor diversity.
How does mIRN-21 micro RNA lead to tumor suppression?
mIRN-21 is a cytoplasmic micro RNA (72 bp) originally transcribed from the an intron of the vacuole membrane protein gene. The product is a stem-loop precursor mRNA which is cut by the endonuclease Drosha, exported to the cytosol, cut again by Dicer and its other strand is then degraded. The final micro RNA targets tumor suppressor genes (3’ UTR) such as PTEN, Bcl2, IL-12p35, and many others.
What part of the endogenous retrovirus genome is capable of altering host gene expression?
Long terminal repeats are at both ends of endogenous retroviruses and serve as viral integration sites into new areas of the genome. They frequently contain promoters, and enhancers capable of influencing transcription of nearby host genes.
How does Sirtuin 1 act on chromatin?
Sirtuin 1 is an NAD-dependent deacetylase related to the silent mating type information regulation 2 homolog 1 of S. cerevisiae. It reverses epigenetic DNA changes favoring gene expression-histone acetylation thereby reducing gene transcription. It may also silence LINES thereby reducing gene hopping induced mutations in the elderly. Low levels in progeria
What is the difference between prokaryotic and eukaryotic gene transcription?
Prokaryotic transcription uses a 5 unit polymerase, transcription sequence are located near the promotor, starts with a ribosomal binding site, formyl methionine loads the ribosome, can begin translation during RNA transcription, can transcribe at multiple sites along the mRNA. Eukaryotic transcription requires nucleosome unwinding, promotor attachment, usually has an untranslated initial sequence, a methionine start site which extend the 5’ UTR, requires different polymerases for different types of genes, needs mRNA splicing to remove introns from primary transcript, attaches a 5’ methylated cap and a 3’ poly A tail, cannot begin translation until transcription and splicing are complete, only one ribosome may attach at a time, the stop codon extends to 3’ UTR.
What controls the progression from one cell stage G1 to S?
Satisfaction of checkpoints and progression of cyclin-dependent kinase (CDK 1-4) attached to various cyclins (a-E) which phosphorylates Rb protein (protein product of retinoblastoma) component of the Rb-E2F complex. Phosphorylation frees E2F transcription factors to express the genes necessary for DNA synthesis (DNA polymerase, thymidine kinase, and dihydrofolate reductase). Phosphorylation is inhibited if DNA has been damaged.
How did Mendel develop the plant material on which to propose his laws of heredity?
He started by breeding plants that consistently had progeny in which all members showed the same trait (homozygous). By crossing these plants with others of different traits the F1 generation showed mixed results in a consistent pattern. He noted which traits were dominant or recessive and independent assortment when studying multiple traits. Using homozygous parents who developed heterozygotes F1 generation necessarily had fixed proportion of different traits.
How are dominant and recessive traits expressed in family trees?
Dominant traits always include one affected parent, showing a vertical pattern of inheritance. Two affected parents, and heterozygotes can occasionally have a normal child. Recessive traits are expressed in all children of affected parents. Horizontal pattern develops with affected children belonging to the same generation. Vertical inheritance pattern occurs if the recessive trait is common in that population. Affected children need not have affected parents but are more common in consanguineous unions.
How have Mendel’s laws been adjusted?
Dominance needed adjustment for degree of dominance. Incomplete-1:2:1 red:pink: white Codominance produces phenotypes 1:2:1. peas- spotted:spotted/dotted:dotted 2->multiple alleles produce phenotypes in a series of 3:1 gain-of-function alleles that are hypermorphic (achondroplasia-over expression of FGFR3 that activates TK and stops bone growth ) , neomorphic (huntington), or antimorphic (antenna to leg). Some recessive alleles are lethal ->2:1 ratios. Some genes influence several traits-pleiotrophy. Genes vary in expressively (degree to which a specific genotype is expressed in a phenotype), and penetrance (degree to which those with the same genotype show the same phenotype).
How may 2 genes interact in a dihybrid F1 cross in the F2 generation?
4 distinct phenotypes-9:3:3:1 seed coar color peas-brown, tan, gray, green. 4x4 table AABB->aabb with 16 cells Complementary: one dominant allele of each gene necessary-9:7 sweet pea flower color purple:white. Recessive epistasis: homozygous recessive blocks both alleles of 2nd gene 9:3:4 labs black brown yellow dominant epistasis type 2: dominant allele B (prevent pigment deposition, b-wild type deposits color) hides both alleles of A gene 12:3:1 squash color-white:yellow:green. green=bb if no A (white) redundancy: only 1 dominant allele of either gene produces phenotype 15:1 Maize leaf
How are open reading frames discovered within genomic DNA?
Isolated clones of genomic dsDNA are searched in both directions 2x3 transcripts for stop codons. Any sequence in any direction that codes for 7 aa in a row without a stop codon is an open reading frame.
What limits the use of cDNA as a way to find genes?
cDNA needs to come from tissues that express the gene of interest which depends on the tissue studied and its metabolic activity.
