Genome to function

T/F: If a female human has a recessive mutation only on one X chromosome, it cannot induce a phenotype

False: if “normal” chromosome X is preferentially inactivated in enough cells, the cells will express the mutant, this can produce a phenotype

T/F: Gene promoters but not enhancers are easy to identify from genomic sequences

T: enhancers are often thousands of bases away from the genes, they dont have universal sequences or motifs. Promoters have recognizeble sequence features

T/F:Intron sequences are translated into peptides and then removed to produce the mature protein sequence.

False

T/F Intronic sequences cannot be identified by computational analysis of genome sequences.

False

T/F Transcription factors commonly have a DNA binding domain

True

T/F For routine sequencing, NGS technology is the first choice.

False

T/F Primers are required for Sanger sequencing

True

Which sequencing uses deoxynucleotides and sorts DNA reads by length

Sanger

T/F: NGS and Sanger sequencing technologies are both based on in vitro DNA replication.

True

What’s an enhancer?

A regulatory sequence of DNA that regulates gene expression by allowing the bind of transcription factors, increasing transcription rate of genes, usually they are far away from the genes that they regulate

What is a promoter?

Sequence of DNA upstream the gene, where DNA polymerase binds

Describe the changes preformed to a non matured mRNA

3’ polyadenilated tail and 5’ methylguanosine cap, protecting mRNA from degradation

Name the histones that compound the nucleosome

8 histones, 2 of each: H2A, H2B, H3 and H4

What is the role of H1

Acts as an stabilizer between each nucleosome

Histones are rich in which amino acids?

Lysine and arginine: net positive charge

What is the charge of DNA

Negative

What is the effect of acetylation in lysine amino acids in histones

It neutralazes the positive histone charge, lowering the interaction with DNA

Name starting codon MET

AUG

Name stop codons

UAA UAG UGA

Aneuploidy

Alterations in the number of chromosomes

Euploidy

Alteration in the number of sets of chromosomes

Chromosome Structural mutation that affect the number of genes

Duplication and deletion

Chromosome Structural mutation that affect the position of genes

Translocations and inversions

Chromosome numeric mutation (affects number of chromosomes)

Aneuploidy: losing or wining one or many chromosomes, Euploidy: gaining one or more sets of cromosomes

Name the four effects of mutations

Neutral (silent), nonsense, nonsynonymous and frameshift

What is a neutral mutation?

A mutation that is produce by the change of a base that still codes for the same aminoacid (redundant genetic code)

What is a nonsense mutation?

A mutation that produces an early stop codon

What is a nonsynonymous mutation

a mutation that changes the encoded aminoacid, usually changes in the 3rd base of the codon generate this kind of mutation.

Frameshift mutation

A mutation that inserts or delites a base, altering the frame lecture of the sequennce, often resulting in a premature stop codon or a different protein

T/F: Genetic code is redundant but some codons are more favored than others

True

T/F: a change for a synonymous codon can reduce gene expression

True

T/F: Having preferences between codons is part of gene regulation

True

What is NMD (Nonesense-mediated decay)?

A surveillance mechanism for errors in gene expression: degradates mRNA with premature stop codons (nonsense mutations)

What are EJCs (Exon-junction complexes)?

Group of proteins that bind the mRNA after splicing in the unions where introns where removed. They direct mRNA export, translation and quality control.

Explain NMD (Nonsense mediated decay)

After splicing and during the translation of mRNA, if the ribosome encounters a STOP codon, it stalls and forms a SURF complex with the EJC, promoting mRNA decay.

T/F: NMD does not need the presence of a EJC to mediate mRNA decay

F: Because EJC-dependent NMD requires a downstream EJC, it is only effective in the coding regions upstream of the last EJC (indicated in blue). It cannot detect any premature stop codons downstream of the last EJC (red part)

T/F: if a premature stop codon in after the last EJC, the NMD can still go on

False: the mechanism always needs a EJC to generate the SURF complex and mediate decay

T/F: Somatic mutation rate is higher than in germ cells

True

Name the effects of mutations

Loss of function (LOF), gain of function (GOF) and Dominant negatives (DN)

What are the types of loss of function mutation effects (LOF)

Hypomorphs, Amorphs and protein null

Hypomorph

LOF mutation effect that generates a partial loss of function

Amorph

A LOF effect that causes complete loss of function

Protein null

A LOF effect where there is no protein at all

T/F: LOF effects often cause recessive effects, but they can be haploinsufficient (loss of one allele can cause a phenotype)

True

Name GOF (Gain if function) mutation effects

Hipermorphs and Neomorphs

What are GOF hipermorphs?

Gain of function hipermorphs, either the gene activity is increased, or the protein expression or protein activity.

T/F: GOF hipermorphs often produce recesive phenotypes

False: GOF hipermorphs produce dominant phenotypes

What are GOF neomorphs

GOFs that causes a novel gene product functions

What are dominant negative antimorphs

Mutation effect that causes the blocking or inhibation of the normal gene product.

T/F: in a negative dominat antimorphs, the gene product competes and antagonizes the function of the normal alele

True

Transgenesis

The process of introducing a transgene into an organism

Transgene

A trangene is any exogenous genetic sequence either from the same species, a different or an atificial sequence.

Name transgenic connstructs

Plasmid, cosmid, BACs (bacterial artificial chromosome) and YACKs (yeast artificial chromosome)

T/F: plasmid integration occurs by regular baterial transformation

True

Name plasmid benefits

Easy to work with, self replicated (endless of numbers of copies), Stable-store

T/F: For large sequences is usfull using plasmids

False

T/F: A Cosmid can transport larger DNA sequences than the plasmid

True

What are the “cos” sites in Cosmids?

The cos sites come from lambda phage, they are “sticky“ (cohesive) sites that allows DNA to be package in the phages and during bacterial infection (trasudcion) it allows the DNA to cirulirize inside the bacteria.

T/F: Resistance genes are ofthen inserted in plasmids and cosmids

True: this allows to latter identification of transgenic bacteria by antibiotic treatment

How does plasmids get inseted in baterias?

by a heat shock that produces chanels in the bacterial membrane: bacterial transformation. Not 100% efficient

How does cosmids get inseted in baterias?

By bacterial transducction (phage infection), super efficient

T/F: Bacterial Artificial Chromosomes can accomodate larger DNA fragments

True

T/F: YACs > BACs > cosmids > plasmids in lenght of DNA sequences they can transport

True

Name transgenic technics in mamals

DNA microinjection, DNA microinjection with retrovisuses, tansposones, embryonic stem cells

DNA microinjection

Injection of a pasmid/cosmid or BAC into the pronucleous of a developing zygote eggs to latter transfer those eggs to a foster mother.

DNA microinjcection big disadvantage

It random integrates into the genome by inserting multiple copies in tandem arrays

T/F: DNA microinjection can generate mosaic founders but not integrate itself in all cell types.

F: if the integration ocurrs in one-cell stage (zygote) potentially all cells carry the transgene. if it ocurrs after many cell divisions it generates mosaic founders

DNA microinjection with retrovirus

In this technique, the DNA is not injected in the pronucleous, the virus enters to the cells through the embryonic celular membrane receptors

T/F: DNA microinjection has a low success rate

T: due to its physical injection to the pronucleous

T/F: In DNA microinjection the injected DNA integration is random and inefficient

True

DNA microinjection with retrovirus advantage

It’s integration is random but clean and stable, it integrates a single copy

Big disadvantage from DNA microinjection with retrovirus

it can be asily silence because of virus-detection machinery (methilation of the integrated DNA)

Transposons for transgenics

Binary mechanism: they require the target sequence (with ITRs that act as the "recognition sites") and a specific transposase protein that cuts and insrtes the sequence in the host genome

T/F: Transposons do not random insert

False

Embrionic Stem Cells

Are derived from early embrios (inner cell mass) and can be cultured to later integrate transgenes. The resulting transgenic ESC can be injected in a zygote or early embryo to generate chimeric offspring.

Homologous recombination for transgenics = gene targeting

it needs a targeting vector that contains: the transgene, homologous sites arround the transgene and an integrase, the recombinase enzimes cut the host genome and the donor DNA vector and then the integrase incorporates the gene to the targeted site.

How to controle transgenic expression?

(Space controls): With gene enhancers, binary gene expression system (Time control): drug system such a Tet-Offs

Time control system Tet-Off

It allows you to turn “off/on“ a transgene by the tetraciclyne presence or abscence

Gene terapy

Treatments using transgenes

AAV (Adeno-associated viruses)

It uses modifided harmless virus to deliver healthy DNA into patients cells to treat genetic diseases. The viral vector enters the cell and realises DNA so that the body produces missing functional proteins.

T/F: AVV treatments work for all cell types, specialy the ones with higher renewal cell rate

F: Since the DNA does not integrates in the host genome, the effect gets lost in each replication cycle, this is why this terapy works better in stable/low renew cells such as neurons, hepatocytes, etc.

T/F: AVV can work with a single administration

True

T/F: In AVV the transgene is inserted in the host DNA

False

Monogenetic trait

A trait that depends only on the expression of a single gene (only one gene produces the phenotype)

Polygenetic trait

Many genes controlle a certain trait

What are indels?

Genetic mutation that combines incertions (in) and deletions (del) of bases, generating changes in the lecture frame: frameshift or in-frame

in-frame mutations

When there is an addition or deletion of multiples of 3, that add or delete aminoacids

What is an haplotype?

Is a physical grouping of DNA variants (polymorphisms) on a chromosome, that tend to be inherited together because they are close to each other on the chromosome, and recombinations between these variants are rare. A haplotype can be limited to a single gene or it can be larger and include multiple genes.

What is a CNV: Copy Number Variations

Large chunks of DNA that are repeated or missing in different amounts across individuals

T/F: genetic variants that do no appear in at least 1% of the population are no loger considered polymorphisms but instead rare variants or mutations

True

Difference between transposons and retrotransposons

Transposones: Cut-past system

Retrotransposns: Copy-paste system