Biotechnology

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Author:
rica_ross
ID:
230390
Filename:
Biotechnology
Updated:
2013-08-18 16:17:31
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Biology GRE
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Biology GRE
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  1. Restriction enzymes
    • Special class of proteins that recognize specific  DNA sequence and cleave them
    • Each gets its name from the bacteria strain it is found in
    • Usually cut at a palindrome DNA sequence
    • EX: GAAATC
    •       CTTAAG
  2. Sticky Ends
    • Some enzymes cut asymmetrically and create over hangs
    • Allow DNA from different source to be cut with the same restriction enzyme
    • Permits manufacturing of recombinant DNA
  3. Recombinant DNA
    Combination of genes from different organisms
  4. How to get foreign DNA into plasmid
    • Cleave foreign DNA of interest with same restriction enzyme you cut the plasmid with
    • Plasmid will have hole that foreign DNA will fit into
    • Recombinant plasmids
  5. DNA Vectors
    These plasmids are a way foreign DNA can be introduced into a bacterial cell
  6. Three things DNA vectors allow you to do:
    • Figure out the funcation of unknown gene in the lab easier, by isolating the process and proteins that are made from unknown gene
    • Produce clinically useful substances, EX: put bacterial plasmid with human insulin gene into diabetic and this will supply them with enough insulin
    • Give bacteria new functions, bacteria can be engendered for new functions by giving them a new gene
  7. How to single out bacteria that will pick up plasmids
    • About 1% will pick up plasmid
    • Scientists apply an antibiotic resistant gene to plasmid then expose colony to antibiotic, so only the ones that took up the resistant gene survives
  8. Gel Electrophoresis
    • DNA can be broken into smalled fragments by restriction enzymes and separated by size
    • DNA is place into small well carved into gel substance
    • DNA will migrate across gel when electrical field is applied
    • DNA is negatively charged overall and it gets pulled to the positive end
  9. Agrose
    • Gel which fibers trap DNA as they move across gel
    • Longer pieces travel less far
  10. DNA sequencing: Sanger Method
    Unknown piece of DNA is heated to make it single stranded, creates template to match complementary olglinucleotides
  11. PCR: Polymerase Chain Reaction
    • A way to rapidly amplify DNA so there are many copies
    • Starts with double stranded DNA, free nucleotides, DNA poly, and Primers
    • Heat turns DNA single standed
    • Cooling lets primers H bond to end of strands
    • DNA poly builds DNA
    • Primers are radioactive or floursently labeled so they show easily on gel
  12. Southern Blot
    • Used to probe DNA for certain sequence via nucleic acid hybridization
    • Design radioactive enzyme with radioactive nucleotides that is complementary to strand you are looking for
    • Cut up with restriction enzyme DNA gel electrophoresis used to migrate ad separate DNA
    • Alkaline solution goes on gel to denature DNA
    • DNA transfered to nylon membrane and binds and glows
    • *great means of finding small amounts of DNA to work with from a larger sample
  13. RFLP: Restriction fragment length polymorphs
    • Restriction enzymes cut DNS at very specific spot
    • If same enzyme is used on two family members the end up DNA piece may be similar in length but no exact depending on slight person to person changes
    • RFLPS are difference in lengths
    • Used to compare DNA across species and make a DNA fingerprint (use in crime scenes)
  14. Cloning
    • Used an electric pulse
    • a cell from a mammary glad of donor fused with unfertilized egg
    • This egg has no nucleus because it was removed with a small needle
    • the nucleus from the donor cell is the new nucleus
    • The new nucleus is diploid (the original was haploid)
    • New cell implanted into surrogate mother
  15. Genomic Library
    • Large collection of bacterial colonies
    • Each containing copies of a particular foreign DNA segment
    • Libraries are constructed from cDNA (complementary DNA) and cloning cDNA into bacterial cells that copy rapidly
  16. cDNA
    • Complementary DNA
    • Created from mRNA by using enzyme reverse transcriptase to make double stranded cDNA from mRNA transcript
    • Has no introns because made from mRNA
  17. Reporter Genes
    • Some genes don't produce that much mRNA or protein even if they are active
    • So to ensue that mRNA is producing protein they attach a reporter gene
    • Reporter genes are at the promoter region and produce copious amount of measurable proteins
    • Also allow scientists to introduce mutations into promoter regions and see effects of mutations

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