(oct 15) Genomics and RNAi: Sequencing, Genome Evolution, and RdDM

What components make up an organism's genome?

The complete genetic material of an organism, including both coding and noncoding DNA regions.

Approximately what percentage of the human genome consists of protein-coding sequences?

1% to 2%

If a species has a diploid count of $2n = 12$, how many chromosomes are present in a normal haploid gamete?

6 chromosomes ($n = 6$)

What chemical feature of a dideoxynucleotide (ddNTP) causes chain termination during Sanger sequencing?

The absence of a 3'-hydroxyl ($-OH$) group on the sugar ring, which prevents the attachment of subsequent nucleotides.

Why does Illumina sequencing accuracy decrease as read length increases?

Individual DNA molecules within a cluster gradually lose synchrony over successive cycles, leading to a mixed fluorescence signal.

Which sequencing approach is best suited for resolving long repetitive genomic regions?

Third-generation long-read sequencing platforms that can span the entire repetitive region and anchor in unique flanking sequences.

What biological phenomenon accounts for the C-value paradox, where genome size does not correlate with organismal complexity?

Variation in the accumulation of repetitive and noncoding DNA sequences, particularly retrotransposons.

How do retrotransposons differ from DNA transposons in their mode of transposition?

Retrotransposons move via an RNA intermediate that is reverse-transcribed into DNA ("copy-and-paste"), whereas DNA transposons move directly via a DNA intermediate ("cut-and-paste").

What is the primary effect of heavy promoter DNA methylation on gene transcription?

Transcription decreases because methylation physically blocks transcription factors and recruits methyl-binding proteins that compact chromatin into heterochromatin.

What is synteny?

The conservation of gene order along a chromosomal block between different species inherited from a common ancestor.

Why does a single syntenic genomic region in rice correspond to two syntenic regions in maize?

The maize evolutionary lineage underwent an ancient whole-genome duplication event after its divergence from rice.

What is the key functional difference between orthologous and paralogous genes?

Orthologs are homologous genes in different species separated by a speciation event; paralogs are homologous genes within the same species separated by a duplication event.

What is a Robertsonian translocation?

A chromosomal fusion of two acrocentric chromosomes near their centromeres, accompanied by the loss of their small short arms.

Which chromosomal event explains the difference in chromosome count between Przewalski's horse ($2n = 66$) and the domestic horse ($2n = 64$)?

A Robertsonian translocation that fused two acrocentric chromosomes to form chromosome ECA5 in domestic horses.

In comparisons of the maize $bz1$ locus between inbred lines McC and B73, what major structural variation was discovered in gene content?

4 out of 10 genes present in the McC strain were completely absent from the B73 genome, demonstrating gene content non-conservation within the same species.

Why is meiotic recombination completely prevented inside a hemizygous genomic region?

Recombination requires homologous chromosomes to align along matching sequences; a hemizygous region lacks a sequence partner on the homologous chromosome.

By what mechanism do Helitron transposable elements move and alter genomic structure?

Rolling circle transposition, which often captures fragments of host genes and redistributes them to new positions.

How does localized DNA methylation influence meiotic recombination frequency?

It suppresses recombination because densely packed, methylated chromatin blocks access to the machinery that initiates crossing over.

In plant breeding, what is linkage drag?

The inheritance of unwanted wild traits alongside a beneficial gene due to reduced recombination in surrounding divergent sequence blocks.

How does Marker-Assisted Selection (MAS) accelerate plant breeding efficiency?

It allows breeders to screen young seedlings using molecular DNA markers instead of growing every plant to maturity to measure phenotypic traits.

What is Copy Number Variation (CNV)?

Differences between individuals in the number of copies of a specific genomic DNA segment, which alters phenotypic expression through gene dosage.

Which gene copy number variation is primarily responsible for the solid white coat phenotype in Large White pigs?

Duplication of a large genomic segment containing the entire $KIT$ gene.

What structural rearrangement of the $KIT$ gene locus produces colour sidedness in cattle?

Translocation of a $KIT$-containing segment from chromosome 6 to chromosome 29 via a circular DNA intermediate.

What proportion of Neanderthal DNA is carried in the genome of an individual modern human of non-African ancestry?

Approximately 2% (generally falling between 1% and 4%).

What estimated percentage of the original Neanderthal genome survives collectively across all living modern humans combined?

Approximately 20% to 33% (one-third).

Which modern human populations exhibit the highest proportion of Denisovan ancestry (~4% to 6%)?

Populations in Oceania, including Papua New Guinea, Australia, and Melanesia.

Genomic analysis showed that the eastern wolf is an evolutionary admixture of which two parent species?

Grey wolf (~75%) and coyote (~25%).

What genomic ratio of parent ancestral species characterizes the red wolf?

Approximately 25% grey wolf ancestry and 75% coyote ancestry.

How does regulatory RNA modify our understanding of the classic Central Dogma of molecular biology?

It shows that many RNA molecules are non-coding regulatory active agents that direct mRNA destruction, block translation, or methylate DNA rather than acting merely as passive messengers.

What is the typical nucleotide length of small regulatory RNAs?

20 to 31 nucleotides.

By what mechanism do small RNAs guide silencing protein complexes to target sequences?

Complementary sequence base pairing.

What are the primary characteristics and biological functions of short interfering RNAs (siRNAs)?

They are 21â€"24 nt long, derived from long double-stranded RNA, and function in antiviral defence and transposon silencing in plants and animals.

What precursor structure gives rise to mature microRNAs (miRNAs)?

Single-stranded transcripts expressed from dedicated MIR genes that fold into hairpin stem-loop structures.

Which class of small RNAs is 26â€"31 nt long, found exclusively in animals, processed without Dicer, and responsible for silencing germline transposable elements?

PIWI-interacting RNAs (piRNAs).

What distinguishes the trigger precursor of a microRNA from that of a short interfering RNA?

A microRNA originates from a dedicated single-stranded genomic MIR transcript folded into a hairpin, whereas an siRNA originates from long double-stranded RNA.

What outcome typically occurs when a small RNA guided by RISC matches its target mRNA with perfect complementarity?

The Argonaute protein acts as an endonuclease and cleaves the target mRNA, leading to complete degradation.

What outcome typically occurs when an animal microRNA matches its target mRNA with imperfect complementarity?

The RISC complex remains bound to the target, blocking ribosome translation and causing gradual transcript degradation.

What structural feature of a primary MIR gene transcript allows processing enzymes like Dicer to act upon it?

Internal sequence complementarity that causes the single-stranded RNA to fold into a double-stranded hairpin stem.

How is one strand of a small RNA duplex selected to become the active guide strand inside Argonaute?

Strand selection is governed by the relative thermodynamic stability of the two duplex ends.

Why is double-stranded RNA (dsRNA) a reliable signal for host cell antiviral defence systems?

Long dsRNA is largely absent from normal uninfected eukaryotic cells, making its presence a clear indicator of viral replication.

How does the RNAi pathway generate sequence specificity against a novel virus without prior host immunity?

Dicer cuts the viral dsRNA itself into siRNAs, using the pathogen's own genetic material to program RISC complexes against viral transcripts.

What evolutionary evidence demonstrates the critical role of RNAi in plant and insect antiviral defence?

Viruses infecting these organisms have evolved specialized viral suppressor proteins designed specifically to disrupt the host RNAi machinery.

How does RNA interference differ from the vertebrate adaptive immune system in foreign target recognition?

RNAi operates intracellularly by recognizing foreign sequence complementary nucleic acids, whereas adaptive immunity relies on antibodies and specialized cells targeting foreign proteins.

What is the primary function of RNA-directed DNA Methylation (RdDM) in plants?

To direct cytosine DNA methylation and long-term epigenetic gene silencing onto target genomic loci using small RNA guides.

Which plant-specific RNA polymerase initiates the RdDM pathway by transcribing repetitive target loci?

RNA Polymerase IV.

What enzyme converts single-stranded RNA Polymerase IV transcripts into double-stranded RNA during plant RdDM?

RNA-dependent RNA polymerase 2 (RDR2).

Which specific Dicer enzyme processes dsRNA into 24-nucleotide fragments in the plant RdDM pathway?

DCL3 (Dicer-like 3).

Which Argonaute protein binds 24-nucleotide small RNAs in plant RNA-directed DNA Methylation?

AGO4.

What is the key functional role of RNA Polymerase V in the RdDM pathway?

It transcribes a non-translated scaffold RNA at the target locus that anchors the small RNA-loaded AGO4 complex via base pairing.

Which DNA methyltransferase enzyme deposits methyl groups onto cytosines at the end of the RdDM pathway?

DRM2.

Why is the RdDM pathway described as self-correcting for transposon control?

Active transcription of a transposon generates the exact small RNA material required to target its own DNA locus for methylation and silencing.

What surprising phenotypic result was observed in Richard Jorgensen's petunia overexpression experiment?

Introducing extra copies of the chalcone synthase gene yielded white or variegated flowers rather than deeper purple flowers, revealing co-suppression.

What key molecular trigger of RNA interference was identified by Andrew Fire and Craig Mello in 1998?

Double-stranded RNA (dsRNA).

Where are MIR genes and microRNAs predominantly localized along plant chromosomes such as in tomato?

In gene-rich euchromatic regions near the distal ends of chromosome arms.

Why are MIR genes concentrated in euchromatic distal chromosomal regions rather than pericentromeric regions?

MIR genes require open, accessible chromatin for active transcription; placement in methylated heterochromatin would silence them.

Where are 24-nucleotide silencing siRNAs predominantly concentrated along plant chromosomes?

In repeat-dense, pericentromeric heterochromatin regions where transposable elements are concentrated.

What defines an acrocentric chromosome?

A chromosome whose centromere sits near one end, producing a very short p arm and a long q arm.

How do whole-genome duplication and domestication differ in maize evolutionary history?

Whole-genome duplication was an ancient natural event millions of years ago, whereas domestication transformed teosinte into modern maize over ~9,000 years via human selection.

What is the transcriptome?

The complete set of RNA transcripts present in a cell at a given moment in time, which varies dynamically with conditions.

Why is the ratio of genes to total sequence content significantly lower in bread wheat compared to humans?

Bread wheat is hexaploid and its genome is heavily inflated by massive amounts of noncoding repetitive DNA sequences.