9.5 Finding the Genes in Genomes

Question 1

A long open reading frame may be part of a _______________

Answer 1

protein-coding exon

Question 2

How many reading frames can DNA be read in?

Answer 2

six

Question 3

What is an open-reading rame (ORF)?

Answer 3

It is a reading frame that is uninterrupted by stop codons
OR
Stretches of nucleotides that have a reading frame of triplet uninterrupted by a stop codon

Question 4

Describe how scientists predict the location of genes by
identifying sequences conserved in the genomes of widely
divergent species. (name of process)

Answer 4

comparative genomics

Question 5

What do the branch points on a diagram depicting the species relatedness genome conservation between H.sapiens and other vertebrates represent?

Answer 5

They represent a series of nested common ancestors. The number of each branchpoint is millions of years before the present.

Question 6

What does it mean when DNA sequences are conserved in highly divergent species?

Answer 6

it means that these DNA sequences may be part of genes

Question 7

What is the percentage in which the human and zebrafish genomes are conserved (column 1)?

Answer 7

2%

Question 8

Within protein-coding sequences what is the percentage of the conservation of the human and zebrafish gemones (column 2)?

Answer 8

82%

Question 9

What tool can be used to discover cross-species homology?

Answer 9

A computerized visualization tool

Question 10

What is a direct method of finding genes (has to do with transcription)?

Answer 10

Locating transcribed regions

Question 11

What are the two reasons that mRNA is not easily sequenced?

Answer 11

1. It is too rare to purify
2. Sequencing technology is not widely available

Question 12

How can mRNA be made nto cDNA for sequencing?

Answer 12

It is known that retroviruses have RNA genomes, so reverse transcriptase is used to copy mRNA into cDNA.

Question 13

What is reverse transcriptase?

Answer 13

It is an enzyme that synthesizes complementary DNA (cDNA) from an RNA template. This process is called reverse transcription and is the reverse of the usual transcription process, where DNA is transcribed into RNA.

Question 14

What are retroviruses?

Answer 14

It is a type of virus with RNA as its genetic material, and it relies on reverse transcriptase to replicate within a host cell. Reverse transcriptase enzymes, originally discovered in retroviruses, are used to reverse transcribe mRNA to cDNA.

Question 15

Converting RNA transcripts to cDNA(steps):

Answer 15

also refer to tutor slides
1. Obtaining mRNA from red blood cell precursors:
Eukaryotic mRNAs have a poly-A tail at the 3’ end. mRNAs are purified by affinity to oligo(dT)- single-strand DNA fragments of 20 nucleotides made of dT(deoxythymidine) only.
2. Synthesis of hybrid cDNA-mRNA molecule:
In vitro synthesis using reverse transcriptase+ dATP+dGTP+ dTTP+ dCTP. Then primer DNA synthesis using oligo(dT)

Question 16

Thereafter, how is the second DNA strand complementary to the cDNA created?

Answer 16

1. mRNA is digested with RNAse
2. 3’ end of the cDNA folds back and acts as a primer for the 2nd strand synthesis
3. In the presence of dNTPs and DNA polymerase, the first cDNA strand acts as a template for synthesis of the second cDNA strand.
4. Double-stranded cDNA can then be cloned into a plasmid

Question 17

Compare genomic libraries and cDNA libraries from each other

Answer 17

Key Difference: Genomic libraries have all DNA (coding and non-coding), while cDNA libraries include only the coding sequences of expressed genes.

Genomic libraries: Clones represent all regions of DNA equally and show what the genome looks like in the region of each clone.
cDNA libraries: clones reveal which parts of the genome contain information used in making proteins in specific tissues.
It is used to find transcribed regions (genes)