110MT-SP09 - BIMM 110 Molecular Basis of Human Disease I.E....

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1 BIMM 110 Molecular Basis of Human Disease Spring 2009 I.E. Scheffler MIDTERM EXAM There are 9 questions. All answers should be written in the blue book. Leave the first inside page blank for scoring and write a column of numbers from 1 to 9 If you want your blue book returned in the hallway near my office, please sign the waiver on the back of the blue book. ***************************** QUESTIO N 1 (18 points) a. [2] What is the difference between an general endonuclease and a restriction enzyme ? An endonuclease can cleave DNA randomly at any internal position, regardless of the nucleotide sequence. A restriction endonuclease cleaves only at restriction sites. These sites are defined by a specific sequence of 4 – 10 bp; the sequence is normally a palindrome (inverted repeat) with a unique bp in the middle. Restriction enzymes make a cut in each strand; in some cases blunt ends are generated, in other cases the cuts are staggered. b. [2]What do we mean when we talk about “sticky ends” produced by a restriction enzyme? When the cuts are staggered, the two ends consist of short single stranded regions extending 2 or more nucleotides beyond the base paired region. c. [6] Vectors used for cloning in E. coli are typically small circular DNAs with a number of useful and essential elements. What are they and what is their function? An origin of DNA replication in E.coli allows the plasmid to replicate A selectable marker (resistance to an antibiotic) allows the selection of bacteria (colonies) that have been successfully transfected. A cloning site (or multiple cloning site) allows the vector to be linearized, resulting in “sticky ends”; DNA fragments with complementary sticky ends can be cloned into this site. The cloning site must be outside of the selectable gene or origin. d. [5] What is the distinction between a genomic and a cDNA library? A genomic DNA library is a collection of fragments from an entire genome cloned into a suitable vector. A cDNA library is made by converting the population of mRNAs from a given tissue into double stranded DNA fragments; this requires first a reverse transcriptase and then a DNA polymerase. These fragments are then also cloned into a suitable vector. In the cDNA library only the exons of a gene expressed in that tissue are represented. e. [3] The complete reaction mixture for the polymerase chain reaction does not contain one or more of the following:
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2 a) all four deoxynucleoside triphosphates. b) DNA containing the sequence to be amplified. c) D N A ligase . d) heat-stable DNA polymerase. e) oligonucleotide primer(s). f) a restriction enzyme (may be needed for RT-PCR) g) reverse transcriptase h) ATP QUESTIO N 2 (12 points) a) [8] What are the absolutely essential elements of a YAC (yeast artificial chromosome) vector to be used in yeast?
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This note was uploaded on 05/01/2010 for the course BIMM 110 taught by Professor Mcginnis during the Spring '08 term at UCSD.

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110MT-SP09 - BIMM 110 Molecular Basis of Human Disease I.E....

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