Course Notebook: Methods DNA, 1976 - 1984

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Course Notebook: Methods DNA, 1976 - 1984

Source

box: 10 folder: 03-04 (text)

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2 Folders

Description

This is a notebook within a series on Grieder's notes on the methods she used. The notebook begins with a dicussion of a new protocol, Sequenase. It was a step by step protocol for DNA sequencing. It also included instructions for sequencing double-stranded templates. The notebook also conatined a protocol Greider made for Tetrahymena Telomere clones in two different plasmids. The protocol used pBR322 and pUC118, which were restriction enzymes that were inserted in plasmids that were cloned at Bam H1 site at both plasmids. It was a different method for cloning vectors and included with it a subsequence document on the labelling of a hexamer, which is a six oligonucleotide sequence, as a probe. The notebook also included an efficient molecular biology test for Vector Cloning System called Gigapack. Gigapeck was an easy to use frozen extract. One would simply thawed the extract and added to the DNA. The kits also included a sample of control lambda DNA and indicator bacteria. Greider also ordered custom oligonucleotides and custom linker. Within this series of protocols there was an interesting document on Epicurian coli, which seemed to be a play on the model organism bacterium E.coli, for what was referred to as gourmet cloning. The gourmet cloning would provide E.coli for ready use. It also included a drawing of high class individuals eating E. coli that had been inserted into a plasmid. Near the drawing was a sign for a Plasmid Social for $10.00.

The notebook also contained a document from Hanahan on Studies on Transformation of E. coli with plasmids to render DNA uptake more efficiently such that virtually every plasmid molecule that interacts with a sodium or pottasium channel becomes transported across the cell envelope. Also included the enzymes needed for the transformation. J.F. Kirsh, a colleague of Greider's at Berkley, sent a letter entitled Purification of Synethic Oligonucleotides by Thin-Layer Chromatography. In it he described a method based on an article from the journal Science. It detailed materials and procedures of different blotting techniques.

The next notebook in this series of methods and meterials involving DNA, are a list of instructions for Yeast Linear Plasmid DNA prep. It seemed to have been created at the Blackburn Labs for spheroplast preperation. However, Greider didn't think the protocol was necessary to completely degrade the cell wall using spheroplasts. By the end of the end of the procedure, Greider noted that the DNA at this point was about 20% of the linear plasmid and was cleaned enough for cutting with restriction enzyme.

Greider does a large-scale plasmid prep using two different solutions and pellets and re-suspends cells.

The notebook also contained instructions of certain procedures. The procedures included agarose elution, glass beads, and agarose elution. The notebook also contains a book on cleaning and storing DNA with notes by Greider on the side where she provided a different procedure for creating different buffers. Douglas Hanahan and Matthew Meselson, colleagues of Greider's in the Blackburn lab, gave a protocol to Greider regarding High Density Plasmid Screening in 1978. The sreening colonies contained recombinant plasmids by colony hybridization at high density. The essential feature of this protocol was a modification of the standard colony hybridization procedure. It essentially was a method for replica plating small colonies at high density with good fidelity. It required the cells to be grown in nitrocellulose filters. The protocal included within it two techniques. One technique required freezing high density filter. The other was a protocol that was applicable to E. coli strains.

There was also a dicussion of colony hybridization and plaque hybridization which was written by Michael Grunstein from the Molecular Biology Institute at University of California at Los Angeles.

The notebook concludes on a document on preparation of Yeast DNA to harvest and suspend cells and centrifuge the cells. Greider noted that the method was relatively long, but gave clean DNA and recovered high and low DNA equally well.

Scope and Contents

This is a notebook within a series on Grieder's notes on the methods she used. The notebook begins with a dicussion of a new protocol, Sequenase. It was a step by step protocol for DNA sequencing. It also included instructions for sequencing double-stranded templates. The notebook also conatined a protocol Greider made for Tetrahymena Telomere clones in two different plasmids. The protocol used pBR322 and pUC118, which were restriction enzymes that were inserted in plasmids that were cloned at Bam H1 site at both plasmids. It was a different method for cloning vectors and included with it a subsequence document on the labelling of a hexamer, which is a six oligonucleotide sequence, as a probe. The notebook also included an efficient molecular biology test for Vector Cloning System called Gigapack. Gigapeck was an easy to use frozen extract. One would simply thawed the extract and added to the DNA. The kits also included a sample of control lambda DNA and indicator bacteria. Greider also ordered custom oligonucleotides and custom linker. Within this series of protocols there was an interesting document on Epicurian coli, which seemed to be a play on the model organism bacterium E.coli, for what was referred to as gourmet cloning. The gourmet cloning would provide E.coli for ready use. It also included a drawing of high class individuals eating E. coli that had been inserted into a plasmid. Near the drawing was a sign for a Plasmid Social for $10.00.

The notebook also contained a document from Hanahan on Studies on Transformation of E. coli with plasmids to render DNA uptake more efficiently such that virtually every plasmid molecule that interacts with a sodium or pottasium channel becomes transported across the cell envelope. Also included the enzymes needed for the transformation. J.F. Kirsh, a colleague of Greider's at Berkley, sent a letter entitled Purification of Synethic Oligonucleotides by Thin-Layer Chromatography. In it he described a method based on an article from the journal Science. It detailed materials and procedures of different blotting techniques.

The next notebook in this series of methods and meterials involving DNA, are a list of instructions for Yeast Linear Plasmid DNA prep. It seemed to have been created at the Blackburn Labs for spheroplast preperation. However, Greider didn't think the protocol was necessary to completely degrade the cell wall using spheroplasts. By the end of the end of the procedure, Greider noted that the DNA at this point was about 20% of the linear plasmid and was cleaned enough for cutting with restriction enzyme.

Greider does a large-scale plasmid prep using two different solutions and pellets and re-suspends cells.

The notebook also contained instructions of certain procedures. The procedures included agarose elution, glass beads, and agarose elution. The notebook also contains a book on cleaning and storing DNA with notes by Greider on the side where she provided a different procedure for creating different buffers. Douglas Hanahan and Matthew Meselson, colleagues of Greider's in the Blackburn lab, gave a protocol to Greider regarding High Density Plasmid Screening in 1978. The sreening colonies contained recombinant plasmids by colony hybridization at high density. The essential feature of this protocol was a modification of the standard colony hybridization procedure. It essentially was a method for replica plating small colonies at high density with good fidelity. It required the cells to be grown in nitrocellulose filters. The protocal included within it two techniques. One technique required freezing high density filter. The other was a protocol that was applicable to E. coli strains.

There was also a dicussion of colony hybridization and plaque hybridization which was written by Michael Grunstein from the Molecular Biology Institute at University of California at Los Angeles.

The notebook concludes on a document on preparation of Yeast DNA to harvest and suspend cells and centrifuge the cells. Greider noted that the method was relatively long, but gave clean DNA and recovered high and low DNA equally well.

Preferred Citation

Course Notebook: Methods DNA, 1976 - 1984. Carol Greider Collection, Cold Spring Harbor Laboratory Archives Digital Repository. 88-1525527. Update 2025-03-18.

Credit Line

Courtesy of Cold Spring Harbor Laboratory Archives.

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Level

file

Collection

Carol Greider Collection

Resource ID

CWG

Reference ID

cc48d7b26683d1c8abdc35016474c864

Finding Aid Link

Course Notebook: Methods DNA