Essential Biochemistry - Texas Tech University

Essential Biochemistry - Texas Tech University

Sanger or Dideoxy DNA Sequencing Components for the Chain Termination Method: DNA fragment for sequencing Primers dNTPs ddNTPs Sanger or Dideoxy DNA Sequencing What direction does the DNA polymerase elongate the DNA fragment?

Sanger or Dideoxy DNA Sequencing Migration smaller is faster through the gel (bottom smallest) Each peak represents 1 base Pyrosequencing Based on DNA synthesis Template DNA is immobilized Peak area

represents base number http://www.pyrosequencing.com/DynPage.aspx?id=7454 Pyrosequencing Potential Problem: DNA polymerase and luciferase compete for the same dATP substrate Solution: Deoxyadenosine alpha-thio

triphosphate (dATPS) is efficiently S) is efficiently used by DNA polymerase but not recognized by luciferase Pyrosequencing Requirement: Generate ATP from polymerase generated pyrophosphate Solution: ATP sulfurylase quantitatively converts PPi to ATP in the presence of adenosine 5

phosphosulfate Pyrosequencing Potential Problem: Unincorporated dNTPs can react in the next cycle producing false positives Solution: The nucleotide degrading enzyme apyrase degrades unincorporated dNTPs

between cycles Gene Amplification by Polymerase Chain Reaction (PCR) Gene specific amplification without purification Amplification via temperature cycling Special Taq (Thermus aquaticus) DNA polymerase Detection of Helicobacter pylori, Borrelia burgdorferi,

as well as hepatitis, HIV, and West Nile virus Restriction Enzyme Recognition Sites Restrictive endonucleases recognize 4-8 bp sequences within the DNA and cleave at a specific site DNA strands that are cut in a symmetric fashion are palindromic

Blunt and Sticky DNA Cuts Molecular Cloning by Recombinant DNA Technology Cut by restriction enzymes Anneal an uncatalyzed reaction Re-close by DNA ligase Screening for Colonies Containing

the Plasmid with the Insert Size-Specific Cloning Vectors Recombinant Protein Products Generating a Restriction Map Where are the restriction sites located on the 20 kb fragment that

would generate this map? . . . Transcript Analysis via Microarray/DNA Chip How to examine gene expression changes with a given treatment

mRNA cDNA fluorecent labeling cDNA/oligonucleotide Hybridization Slide scanning Data analysis Differential Arabidopsis Gene Expression

with GB03 Exposure RNA Extraction cDNA Synthesis Label Hybridize Wash Scan Block 8 Full Image

cDNA Synthesis and Labeling O HN - O O O

O P O P O P O- O- O-

O O O NH2 N OH Amino Allyl-dUTP -

O3S N N O O O O

Cy5 Dye N SO3- Microarray Data Analysis

Recently Viewed Presentations

  • Brisbane Organic Growers

    Brisbane Organic Growers

    Pick up a pH test kit from your garden centre. Do the simple test from various areas in the garden. pH 6-7.5 neutral, suits most plants, maximise nutrient availability. pH < 5 acidic, most plants will not grow well and...
  • ISMS implementation overview seminar - ISO27001security

    ISMS implementation overview seminar - ISO27001security

    The RTP should be set within the context of the organization's information security policy and should clearly identify the approach to risk and the criteria for accepting risk. The RTP is the key document that links all four phases of...
  • NURSES SERVICE, RANK, NAME Insert your photo here:

    NURSES SERVICE, RANK, NAME Insert your photo here:

    I knew I wanted to be a nurse when . I was a 20-year-old undergraduate student and saw the professionalism and dedication of the Air Force nurses caring for my closest friend, who experienced severe complications after giving birth. When...
  • Bones and Skeletal System - SCIENTIST CINDY

    Bones and Skeletal System - SCIENTIST CINDY

    Between the diaphysis and each epiphysis of an adult long bone is an epiphyseal line. This line is a remnant of the epiphyseal plate, commonly called the . growth plate, a disc of hyaline cartilage that grows during childhood to...
  • WedNesday May 7th 2014 QU: What is a

    WedNesday May 7th 2014 QU: What is a

    Thursday May 8th 2014. QU: Keeping a food web in mind, why would a more diverse ecosystem be better? OBJ: Ch15 Notes, Fluctuations of a Food Web,
  • Selection Process - Colorado State University

    Selection Process - Colorado State University

    Digital collections created from the Libraries' archival holdings to support teaching and research. Select archival materials, generated by CSU entities such colleges and departments, that document the history and intellectual life of CSU ... fluid, and grows in a ....
  • The Metric System

    The Metric System

    The last digit of any measured number is assumed to be an estimate (uncertain) The second to last digit is assumed to be known with certainty (based on a line) A (25.5 mL) B (16.37 mL) C (51 mL) 26....
  • History of Film - Amazon S3

    History of Film - Amazon S3

    The Phenakistoscope, a spinning wheel with an image at it's center that seems to move, is invented by Joseph Plateau in Belgium. ... Charlie Chaplin's tramp character first appears in . Kid Auto Races at Venice. Lois Weber's feature-length parable...