Biochemistry 2280A Lecture : Constructing a Genomic Library

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orchidotter238

29 Dec 2011

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Isolated from many cells human dna cleave with restriction nuclease. Gram negative bacteria genome ~ 2 million bp. The h. influenze genome is 2 million bp. 1000 (2 000 000/2000) = 1000 would be the absolute minimum. To be sure you have full coverage you would want an excess of several fold. Start with millions of cells extra dna, sonicate it to get fragments dna fragments of various sizes. Use dna to make the genomic library agarose gel electrophoresis. 20 000 clones; each represents an independent fragment of the genome. One close: isolate plasmid dna, anneal prier, dideoxy sequence. Sequence the ends of all 20 000 genomic clones; obtain end-sequences of dna insert; end-sequences. 25 000 sequence runs from 20 000 clones. Put 25 000 sequences together in the correct order. A contiguous dna sequence representing a portion of the genome (assembled from many independent sequences much bigger than 1 clone)

Related Questions

DNA libraries are useful for identifying and isolating eukaryotic DNA fragments of interest in research. Select the ways in which genomic and cDNA libraries differ.

1. Genomic libraries contain DNA fragments from an organism\'s entire genome.

2. Genomic libraries are difficult to express in prokaryotic cells, which lack a splicing mechanism.

3. cDNA libraries contain more sequence information than genomic libraries.

4. cDNA libraries are the DNA fragment collections that are stored and propagated in host cells through cloning.

We have three plasmids: the cloning vector, pGEM, the one thatresult if BamHI had been used to construct the genomic library, andthe one that would result if EcoRI had been used to construct thelibrary. What is the expected number of DNA fragments that wouldresult from digesting...

pGEM with the restriction enzymes BamHI, EcoRI, and BamHI +EcoRI?

pGEM + BamHI insert with the restriction enzymesBamHI, EcoRI,and BamHI + EcoRI?

pGEM + EcoRI insert with the restriction enzymesBamHI, EcoRI,and BamHI + EcoRI?

silverotter963

Are the following statements true or false? Please explain your answers.

1. Digestion of genomic DNA with AluI, a restriction enzyme that recognizes a four-nucleotide sequence, produces fragments that are all exactly 256 nucleotides in length.

2. To make a cDNA library, both a DNA polymerase and a reverse transcriptase must be used.

3. It is possible for a coding region of a gene to be present in a genomic library prepared from a particular tissue but to be absent from a cDNA library prepared from the same tissue.

Biochemistry 2280A Lecture : Constructing a Genomic Library

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