Re: [問題] BRA~~BRA

[BLACKOA (怎麼會孤單)]

Outline how would you sequence the gene you cloned in pBR322 by Sanger's method
Please indicate the enzymes and reagents required for sequencing.(10&)

A recomninant plasmid was constructed by inserting an intact human β-globin
cDNA into a restriction endonuciease-cleavage site in pBR322. The recombinant
plasmid was introduced into E.coli and selected. However, β-globin protein was
not expressed in the clone containing full length of β-globin cDNA. Why?

λgt10 is a vector for cDNA cloning. It contains a single EcoRI cleavage site
within the phage repressor gene. Phage libraries cloned in λgt10 are useful
for screening with nucleic acid probe. Explain how cDNAs are synthesized and
cloned in this vector for cDNA library construction. Also explain the
fundamental basis for knowing whether the vector contains cDNA insert in this
strategy(Hint:the control of lytic cycle mechanism.)

(T or F)
1.Both DNA polymerase I and RNase H are required for the synthesis of second
  strand cDNA.

2.DNA methylation is involved in regulation of eukaryotic gene expression.

3.A relaxed plasmid is maintained as a single copy in a cell.

4.σ factor of prokaryotic RNA polymerase II is inhibited by α-amnitin.

5.σ factor of prokaryotic RNA polymerase II must be present for transcription
  to occur.


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