BMSC 210 - Part 5

tasnimjaisee's version from 2017-11-22 05:50

Section Biotechnology

Question Answer
Is REases widespread in prokaryote or eukaryotes?pro
Cuts phosphodiester backbone resulting in double strandREases
Which type of REases is most useful for specific DNA manipulation?ll
Which type of REases was the first REases discovered and purified?l
Which type of REases cuts DNA at random far from their recognition sequence?l
Which type of REases no discrete restriction fragments or distinct gel-banding patterns produced?l
Which type of REases is large; combination restriction and modification enzymeslll
Which type of REases cleaves outside of recognition sequences (~25 bp from site)lll
Which type of REases requires two such sequences in opposite orientations within the same DNA molecule for cleavagelll
Which type of REases rarely give complete digestslll
Structure of cleaved product may differ as?3' or 5' overhang and blunt ends
recognize inverted repeat sequencesPalindromes
EcoR1: Creates ___ nucleotide sticky ends with complementary ___’ end overhangs of ____4 - 5 - AATT
EcoR1: The nucleic acid sequence where the enzyme cuts is?GAATTC
EcoR1: The nucleic acid complementary sequence is _____CTTAAG
What does ligation by DNA ligase do?rejoin two sugar-phosphate backbones of DNA through covalent binding
What are each restriction enzyme partnered with?corresponding modification enzyme sharing SAME recognition sequence
Modification generally consists of _____ of DNAmethylation
After modification, can DNA still be cut by RE?No
Three main steps of gene cloningFragmentation - Insertation - Introduction
_____ are natural vectors and have useful properties as cloning vectorsPlasmids
___ is a common cloning vector, Has all essential elements of a cloning vectorpUC19
pUC19 is a modified ____ plasmidColE1
What is the role of the DNA ligase?Catalyzes joining of 2 strand DNA between 5' & 3'OH
Reverse transcriptaseConverts RNA into DNA
The ___ fragment lacks 5'3' exonuclease activityKlenow
Blue/white screening (bacteria)Bacteria with cloning vector which may or may not contain “insert” DNA
lacZ is inactivated by insertion of ____ ____ and ____ is NOT producedforeign DNA - B-galactosidase
In blue-white Screening, colonies have foriegn DNA inserted?White
Which RNA is used to clone expressed genes?mRNA
How is cDNA made?mRNA used for retroviral reverse transcriptase
Electrophorosis gels contain which chemicals?agarose or polyacrylamide
Steps in PCR amplification Denaturation - Polymerase - Heat and cool
Reverse Transcriptase PCR: Using mRNA stepsAddition of primer & RT - Single-strand cDNA - Add RNase H - Add 5' end and Taq
How can PCR products can be used for cloningJoin PCR product with DNA ligase - Recombinant vector with interrupted lacZ
Segment of single-stranded DNA that is used in hybridization and has a predetermined identitynucleic acid probe
a hybridization procedure where DNA is in the gel and probe is RNA or DNASouthern blot
a hybridization procedure where RNA is in the gelNorthern blot
What kind of probe does souther blot use?Radioactive
What kind of probe does souther blot use?5S RNA
Encode proteins that are easy to visually detect and assayReporter gene
Gene fusionspromoters swapped with reporter genes to elucidate gene regulation under various conditions
Site-directed mutagenesisperformed in vitro and introduces mutations at a precise location
Site-directed Mutagenesis Using Synthetic DNA Oligonucleotides to Create Specific MutationsClone single strand vector - Add oligonucleotide with 1 base mismatch - Extend single strand DNA polymerase
When does Gene disruption occur?Cassettes are inserted into a gene
What does Gene disruption cause?knockout mutations
Gene Disruption by Insertional InactivationCut with EcoRI & ligate - Cut with BamHI & transform into cell with wild-type gene X - Recombination and selection for kanamycin-resistant cells

Section Microbial Genomics

Question Answer
1st generation DNA sequencingSanger Dideoxy Method
DNA replication of a ss template (4)DNA primer initiates complementary strand synthesis - DNA polymerase - 4 dNTPs - ddNTPs
Where do Dideoxy analogs lack –OH groups?At site where another nucleotide attaches to the chain
What does an analog prevent?DNA chain extenstion; resulting in short DNAs
when is Polyacrylamide used?when fragments are shorter
How is a Dideoxy analog formed?Missing OH (replaced with H) compared to normal Deoxyneucleotide
fluorescent labels are detected by who?capillary electrophoresis
Which instrument measures release of light?454 sequencing system
Genome Assembly: what the computer doesCleaves - Find overlap - Deduce
converting raw sequence data into a list of genes present in the genomeAnnotation
Structure of an ORFComputer finds: Start - Stop - Codons between start-stop - Possible RBS - Codon bias - Genuine
Length of virus determined by?length of nucleic acid
Width of virus determined by?size and packaging of protein subunits
Enzymes that cleave glycosidic bondsNeuraminidases