run restriction digestion: ta's will take the pictures for you. review some points sequencing...

triction digestion: TA's will take the pictures for you. some points ing techniques sequencing data

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Run restriction digestion: TA's will take the pictures for you. Review some pointsSequencing techniquesAnalyze sequencing data

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Page 3: Run restriction digestion: TA's will take the pictures for you. Review some points Sequencing techniques Analyze sequencing data

Absorbance (4μl in 400μl water) = 0.05

Concentration = 0.05 * 50 (μg/ml) * 100= 250 μg/ml =250 ng/μl

c1*v1 = c2 * v2

1's = stock solution

c1= 250 ng/μlv1= ?c2 = 6 ng/μl (you needed 300ng in 50 μl= 6 ng/μl)v2 = 50 μl

v1= 6*50/250 μl = 1.2 μl

Calculating the volume of miniprep plasmid DNA solution for your restriction assay

Page 4: Run restriction digestion: TA's will take the pictures for you. Review some points Sequencing techniques Analyze sequencing data

Schematic of the map-based cloning process.

Jander G et al. Plant Physiol. 2002;129:440-450

Page 5: Run restriction digestion: TA's will take the pictures for you. Review some points Sequencing techniques Analyze sequencing data

TAIR worksheet: 8. Which are the genes on either side of GA20OX-1?

Page 6: Run restriction digestion: TA's will take the pictures for you. Review some points Sequencing techniques Analyze sequencing data

TAIR worksheet: 8. Which are the genes on either side of GA20OX-1?

AT4G25420TTA

CAT

ATG

TGA

AT4G25430

AT4G2510

Page 7: Run restriction digestion: TA's will take the pictures for you. Review some points Sequencing techniques Analyze sequencing data

Automated dideoxy DNA sequencing with fluoresncent ddNTPs

http://www.mun.ca/biology/scarr/Fluorescent_didoxy_sequencing.htmlhttp://www.dnassequencing.com/2011/01/07/automated-dna-sequencing-6/

Page 8: Run restriction digestion: TA's will take the pictures for you. Review some points Sequencing techniques Analyze sequencing data

Next generation sequencing: Illumina/Solexa sequencing

Page 9: Run restriction digestion: TA's will take the pictures for you. Review some points Sequencing techniques Analyze sequencing data

Next generation sequencing: Illumina/Solexa sequencing

Advantages: FastCheap (overall)Can be used for RNA sequencing and quantification

Disadvantages: Sequence reads are short

Page 10: Run restriction digestion: TA's will take the pictures for you. Review some points Sequencing techniques Analyze sequencing data

BLAST

The Basic Local Alignment Search Tool (BLAST) finds regions of local similarity between sequences. The program compares nucleotide or protein sequences to sequence databases and calculates the statistical significance of matches.

BLAST can be used to infer functional and evolutionary relationships between sequences as well as help identify members of gene families.

http://blast.ncbi.nlm.nih.gov/blast.cgi

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Ankylosing spondylitis and HLA-B27: polymorphism sequencing › content › annrheumdis › 51 › 7 › 855.full.pdf · Ankylosing spondylitis and HLA-B27: restriction fragment length

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INSTRUCTION MANUAL...minutes. DNA purified using the ZR-96 DCC -5 is suitable for nucleotide sequencing, microarray analysis, PCR, and restriction endonuclease digestion procedures

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