Lecture5mcb160Lf2011

Lecture5mcb160Lf2011 - Dear Students, Slide 2 and 3 are a...

Info iconThis preview shows pages 1–4. Sign up to view the full content.

View Full Document Right Arrow Icon
Dear Students, Slide 2 and 3 are a summary of what Dr. Sundaresan went over at the end of class in Lecture 4. Slide 4-6 are repeats of his last three slides, so you do not need to print out necessarily. However note that I did add some additional information, highlighted in red.
Background image of page 1

Info iconThis preview has intentionally blurred sections. Sign up to view the full version.

View Full DocumentRight Arrow Icon
Summary of Subcloning Experiments Two Subcloning experiments 1) directional cloning using an insert cut with two different restriction enzymes and a vector cut with the same two enzymes (EcoRI-XhoI). Insert: GFP New vector: pGEX-4T This is an example of directional cloning that also allows for in-frame translation of the gfp gene sequences into the GFP protein product. See lab manual p. M-47 for more detail- you will carry out an exercise for this concept later in the quarter. 2) non-directional cloning using a single enzyme Inserts: HindIII digested lambda Vector: HindIII digested Bluescript This is an example of shotgun subcloning in which one adds a collection of insert fragments to prepared vector; can be used to then select transformants and analyze plasmids to determine which bacteria received the fragment of interest, or to generate a library of many fragments.
Background image of page 2
These experiments also illustrate two different methods for improving efFciency of ligating inserts into vector and preventing recircularization of vector (a) directional cloning using a vector cut with two different restriction enzymes and an insert cut with the same two enzymes. If the two enzymes do not have overhangs with complementary DNA sequences, the cut ends of the vector cannot ligate together.
Background image of page 3

Info iconThis preview has intentionally blurred sections. Sign up to view the full version.

View Full DocumentRight Arrow Icon
Image of page 4
This is the end of the preview. Sign up to access the rest of the document.

This note was uploaded on 01/25/2012 for the course MCB 160L160L taught by Professor Venkatesansundaresan during the Fall '11 term at UC Davis.

Page1 / 12

Lecture5mcb160Lf2011 - Dear Students, Slide 2 and 3 are a...

This preview shows document pages 1 - 4. Sign up to view the full document.

View Full Document Right Arrow Icon
Ask a homework question - tutors are online