Biochemistry 2280A Lecture 23: Brandl Topic 23

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magentacrocodile542

20 Feb 2017

School

Western University

Department

Biochemistry

Course

Biochemistry 2280A

Professor

Boffa/ Mc Lauchlin/ Brandl

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Document Summary

Impossible to obtain a sequence of more than 750bp in a single experiment. Standard approach for sequencing small prokaryotic genomes. Does not require any prior knowledge of the genome (no need for genetic or physical map) Data analysis becomes disproportionately more complex as the number of fragment increases. Lead to errors when repetitive regions of a genome are analysed. Genome map provides a guide for the sequencing experiments by showing the positions of genes and other distinctive features. Same approach as the standard shotgun procedure. Uses the distinctive features on the genome map as landmarks to aid assembly of the master sequence. Clone contig method genome is broken into segments (few hundred kilobases/few megabases) which are short enough to be sequenced accurately by the shotgun method usually by partial restriction. These fragments are cloned in a high-capacity vector. A clone contig is built up by identifying clones containing overlapping fragments which are then individually sequenced by the shotgun method.

Related Questions

I just need someone to interpret exactually what I'm suppose todo here. I understand how to compare the sequecnces, however Idon't know exactually how to put them in order. also I can't findout how to find the what chromosome the sequence came from. So farall I gather to do from this is to find which segments overlap, addthe length, tell how big the chromesomeContigs and ShotgunSequencing
In this chapter, we discussed how wholeâgenome shotgun sequencingmethods can be used to assemble chromosome maps. Recall that in thetechnique of shotgun cloning, chromosomal DNA is first digestedwith different restriction enzymes to create a series ofoverlapping DNA fragments called contiguous sequences, or"contigs." The contigs are then subjected to DNA sequencing, afterwhich bioinformaticsâbased programs are used to arrange the contigsin their correct order on the basis of short overlapping sequencesof nucleotides.

In this Exploring Genomics exercise, you will carry out asimulation of contig alignment to help you understand theunderlying logic of this approach to creating sequence maps of achromosome. For this purpose, you will return to the NationalCenter for Biotechnology Information BLAST site that was used inother Exploring Genomics exercises and apply a DNA alignmentprogram called bl2seq.

Exercise I: Arranging Contigs to Create a Chromosome Map
1. Access BLAST from the NCBI Web site athttp://www.ncbi.nlm.nih.gov/BLAST/. Locate and select the "Aligntwo sequences using BLAST (bl2seq)" category at the bottom of theBLAST home page. The bl2seq feature allows you to compare two DNAsequences at a time to check for sequence similarityalignments.

2. Below are eight contig sequences taken from an actual humanchromosome sequence deposited in GenBank. For this exercise we haveused short fragments; however, in reality contigs are usuallyseveral thousand base pairs long. To complete this exercise, copyand paste two sequences into the Align feature of BLAST and thenrun an alignment (by clicking on "Align"). (Hint: Access thesesequences from the Companion Web site so that you can copy andpaste the sequences easily.) Repeat these steps with othercombinations of two sequences to determine which sequences overlap,and then use your findings to create a sequence map that placesoverlapping contigs in their proper order. A few tips toconsider:

â¢Develop a strategy to be sure that you analyze alignments for allpairs of contigs.
â¢Only consider alignment overlaps that show 100 percent sequencesimilarity.
Sequence A
CTAATTTTTTTTGTATTTTTAATAGAGACGAGGTGTCACCATGTTGGACAGGCTGGTCTCGAACTCCTGACCTCAGGTGATCTGCCCACCTCAGCCTCCCAAAGTGCTGGGAT
TACAAGCATGAGCCACCACTCCCAGGCTTTATTTTCTATTTTTTAATTACAGCCATCCTAGTGAATGTGAAGTAGTATCTCACTGAGGTTTTGATTTGCATTTTTCTATGACA
ATGAACAATGTTTCATGTGCTTGTTGGCTGTTTGTATATCCTTTTTGGAGAAATACCAATTCATGTCCTTTGCCCATTTTTAAAGTGGATTGCATGTCTTTTTGTTGTTTAGT
TGTAAAGATGTGGGTTTTTCTTTTGAGACGGAGTCTCGCTGTCGCCTAGGCTGGAGTGCAGAGGCATGATCTCGGCTGACTGCAATCCCCACCTCCTGGCATCAAGAAGTTCT
CCTGCCTCAGCCTTCCAAGTAGCTGGGTTTACAGATGC

Sequence B
CTTTATCTCAGGACAATGAACCCGCAAGGAGAGGAAGAGCCAGTAATTCTATAGAGACTCGGAGGCGCAGGGGGCACGCTTAGTTAGAGTGGTGGTGGTATTTTCAGTGTTTT
CTGGTTTTATGATAAACACAAGCATCAATGTCTCAAGACTTTCATCTTTATCTTTTTTTTTTTTTTTTTTTTTTTCTTGAGACAGGGTTTCCCTCTGTCACCCAGGCTGGAGT
GCATTGGTGGTGTGATCTTGGCTTTCTGTAACCTCGGGCTTCTGGGCTCAAGCCGTTCTACTACCTCAGCCTCCCAAATAGCTAGAACTACAAGCGTGTGCTGCCACACCTGG
CTAATTTGTTGTATTTTATTTATTCATTTATTTTTGTGAAGACAAGGTCTTGCCATGTTGCCCAGGCTGGTCTCAGACTCCTGGGCTCAAGCAATCCACCCACCTTAGTCTCC
CAAAGTGCTGGGATTACAGGCGTGAGCCACCACACCCA

Sequence C
GGAATTTCACTCTTGTTGCCCAAGTTGGAGTGCAATGGCGCGATCTCAGCTCACTGCAACCTCCGCCTCCCAGGTTCAAACGATTCTCCTGCTTCACTCTCCCCAGTAGCTGG
GATTACAGGCTGCACCACCACACCTGGCTAATTTTTTTTGTATTTTTAATAGAGACGAGGTGTCACCATGTTGGACAGGCTGGTCTCGAACTCCTGACCTCAGGTGATCTGCC
CACCTCAGCCTCCCAAAGTGCTGGGATTACAAGCATGAGCCACCACTCCCAGGC

Sequence D
GCTTCATCTTTCTCTTCACCGTAAAACAGGAAAGTGTGTGGTGACCAGTATTTTAAGGGAAAGGCACTTACAGAGAATTAAGCATTTGACAAAATTTATTTACAGATATTTGT
CTGTGGACCACTTCCGCACCAGCTGTGCATGAGAGGGCTCATTGCTCTGAATTTGCCTCCTTGTCTGCACCCAGGAGACCGTTTCCCAGATCACGCAAACGCTGCCTTCTCCC
CACACCAGGGCCCTCAGCATGGGAATGACCTTCCAGCGCTGCACGTTTCCAATCCATGCTCTGTTTTTCAGTTCTGGCTCACAGAGGACTGCTGGTTGCAAGCAAACTTGTAT
CTGGGTCTTCA

Sequence E
CCTTAAGTGATCTACCTGTCTCTGCCTCTCAAAGTGCTGGGATTGCAGGCATAAGCCGCCATGCCCGGCCCAAAGTTTCTTTATATGTGCTGGATACTAGGCCCGTAACAGAT
ATACAATTTGTAAATATTTTCTCTCATTTTGAAGATTTTCTTTTCACTTTCTTGATAATGTCCTTTGTGTATTTTTTGATAATGTCCTTTGATACACAAAAGTTTTTAAGTTT
GATGAAGTTCAATTTACCTATTATTTTCTTTTGTTGTTCAT

Sequence F
TGTTTGTTTGTTTGTTTTGTTTCATTTTGTTTTTGAGACAGAGTCTTGTTCTGTCGCCCAGTGTAGAGTGCAGTGGCATAATCTCGGCTCACCGTAACCTCCGCCTCCCGGGT
TCAAGCAACTCTGCCTGCCTCAGCCTCCCAAGGAGCTGGGATTATAGACGCCCACCACCATGCCTGGTTAATTTTTGTAGTTTTTTTTAGTAGAGATGGGGTTTTGCCATTTT
GGCCAGGCTGGTCTTGAACTCCTGACCTCAGGTGATCTGCCCACCCTGGCCTCTCAAAGTGCTGGGATTACAGGTGTGAGCTGCCACACTCGGCCACAACAAATTTTTGCACC
AGTTGCTCACA

Sequence G
TCCTTTGATACACAAAAGTTTTTAAGTTTGATGAAGTTCAATTTACCTATTATTTTCTTTTGTTGTTCATTCATTTTGTGTCCTATGTAGGAATCTATTGCCAAATTCAAGGT
GATAAAGATTTACCCCTATGTTTCCTTCTAAGAGTTTTATTGTTTTAGCCCTGATATTTAGCTAAACTTAATTGATTTATTAAGTTTAATTTTCCTATGTGGTATGAAGTCAT
TTATCTTCTTTAGTTCAGGATCCAAGTGAAAGGGGCATCTTCTATCTGGGACATGCCATTCTCATGACAGAGGAAAAAGACAAAAAACTGACACATACAATGACTTTAAAACT
TCACTCA

Sequence H
GGGTTTTTCTTTTGAGACGGAGTCTCGCTGTCGCCTAGGCTGGAGTGCAGAGGCATGATCTCGGCTGACTGCAATCCCCACCTCCTGGCATCAAGAAGTTCTCCTGCCTCAGC
CTTCCAAGTAGCTGGGTTTACAGATGCCCACCACCATGCCTGGCTGGTTTTTGTATTTTTAGTAGACACGGGGTTTTACCATGTTGGCCGGGCTGGTCTGGAACTCCTAACCT
TAAGTGATCTACCTGTCTCTGCCTCTCAAAGTGCTGGGATTGCAGGCATAAGCCGCCATGCCCGGCCCAAAGTTTCTTTATATGTGCTGGATACTAGGCCCGTAACAGATATA
CAATTTGTAAA

3. On the basis of your alignment results, answer the followingquestions, referring to the sequences by their letter codes (Athrough H):

a.What is the correct order of overlapping contigs?
b.What is the length, measured in number of nucleotides, of eachsequence overlap between contigs?
c.What is the total size of the chromosome segment that youassembled?
d.Did you find any contigs that do not overlap with any of theothers? Explain.
4. Run a nucleotideânucleotide BLAST search (BLASTn) on any of theoverlapping contigs to determine which chromosome these contigswere taken from, and report your answer.
segment is, and what chromosome it came from. here's theassignment. I just need someone to interpret how I should go aboutdoing this.

viridianox52

QUESTION 1

A mutation caused by exposure to gamma rays is called a spontaneous mutation

	a.	True
	b.	False
	c.	Sometimes, depending on the type of base change
	d.	Gamma rays do not cause mutations

QUESTION 2

A recessive mutation affecting an essential biochemical pathway can be detected________

	a.	when the organism is heterozygous for that mutation.
	b.	when the organism is homozygous for that mutation.
	c.	in either homozygous or heterozygous conditions.
	d.	only by sequencing (recessive mutations never show a phenotype).

QUESTION 3

During DNA replication, the newly synthesized chain grows by adding

	a.	nitrogenous bases
	b.	the sugar-phosphate backbone
	c.	nucleosides
	d.	dNTPs

QUESTION 4

During replication, the DNA strand 5â- AAGTCTAGCCTAG -3â will serve as a template for the polymerization of:

	a.	5â- CTAGGCTAGACTT -3â
	b.	5â - TTCAGATCGGATC -3â
	c.	5â - GATCCGATCTGAA -3â
	d.	3â - AAGTCTAGCCTAG -5â

QUESTIOn 5

Given the following DNA sequences, select which statement(s) is(are) correct.

Sequence 1:

5â-TGGACGCTAA-3â

3â-ACCTGCGATT-5â

Sequence 2:

5â-AATCGCAGGT-3â

3â-TTAGCGTCCA-5â

Sequence 3:

5â-ACCTGCGATT-3â

3â-TGGACGCTAA-5â

	a.	Sequences 1 and 2 are the same
	b.	Sequences 1 and 3 are the same
	c.	Sequences 2 and 3 are the same
	d.	Sequences 1, 2 and 3 are all different

QUESTION 6

Repetitive sequences in the genome are hotspots for:

	a.	Deamination
	b.	Depurination
	c.	Thymine dimer formation
	d.	Replication errors

QUESTION 7

Select which statement(s) about the Ames test is(are) correct

	a.	it has been designed to understand mutation repair systems in Salmonella typhimurium
	b.	it uses mammalian liver extract
	c.	it allows to study whether chemical compounds or their enzymatic breakdown products are mutagenic
	d.	it is based on whether a chemical compound causes reversion mutations from his+ to his-
	e.	two of the above are correct
	f.	three of the above are correct

QUESTION 8

Select which statement(s) is(are) correct.

	a.	All DNA strands have a direction, and it is specified by the carbons in the sugar backbone.
	b.	All DNA strands have a direction, and it is specified by hydrogen bonds between nucleotides.
	c.	In order to form proper base pairs in a double stranded DNA molecule, the two strands must run in opposite directions.
	d.	A and C are correct.
	e.	B and C are correct.

QUESTION 9

Studies of gene mutation frequencies have shown that:

	a.	mutations are rare, and genomes are generally stable.
	b.	mutation frequencies differ among organisms and also between genes, suggesting certain genes are more susceptible to mutation.
	c.	mutation frequencies are consistent between organisms, and each region of DNA is equally susceptible to random mutations.
	d.	Both A and B are correct.
	e.	Both A and C are correct.

QUESTION 10

The compound 5-bromodeoxyuridine (BrdU) is a derivative of uracil, and if BrdU becomes incorporated during DNA replication, it pairs with adenine. This compound is best classified as which type of mutagen?

	a.	base analog
	b.	base inducer
	c.	intercalating agent
	d.	oxidative agent
	e.	alkylating agent

QUESTION 11

The rate of mutation of the fruit fly is higher than the rate of mutation of Algae

True

False

QUESTION 12

Thymine dimers are most commonly caused by which of the following?

	a.	X-rays
	b.	Alkylating agents
	c.	U.V. irradiation
	d.	DNA intercalating agents

QUESTION 13

What chemical group is found in the 3â end of a DNA strand?

	a.	an alcohol
	b.	a hydroxyl
	c.	a methyl
	d.	a phosphate

QUESTION 14

What chemical group is found in the 5â end of a DNA strand?

	a.	an alcohol
	b.	a hydroxyl
	c.	a methyl
	d.	a phosphate

QUESTION 15

Which is the correct order of molecules binding to DNA during DNA replication?

	a.	Helicase, SSB, primase, DNA pol III, DNA pol I, ligase
	b.	SSB, DNA pol I, ligase, helicase, DNA pol I, primase
	c.	primase, helicase, DNA pol III, ligase, DNA pol I, SSB
	d.	SSB, helicase, primase, DNA pol III, DNA pol I, ligase

QUESTION 16

Which statement(s) is(are) correct about strand slippage?

	a.	It can cause disorders such as Huntingon disease.
	b.	It is a process that causes mutations altering the number of DNA repeats.
	c.	It is a process that incorporates nucleotide base analogs and trinucleotide repeats.
	d.	a and b are correct.
	e.	b and c are correct.
	f.	a, b and c are correct.

help needed asap.

chocolatesea-lion434

Biochemistry 2280A Lecture 23: Brandl Topic 23

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