Part D: Lecture 34 Flashcards
Subcloning (def.)
moving an insert from one vector to another
plasmid vector size
1-3 kb
insert DNA for plasmid vector size
< 15 kb
BAC vectors stands for ___
Bacterial artificial chromosomes vectors
BAC vectors size
100 kb E.coli fertility plasmid (sex pilus)
insert DNA for BAC vector size
100 - 300 kb
why make BAC clones?
clone all DNA from an organisms
Making BAC clones procedure
1) isolate genomic DNA
2) shear into large random pieces (100 -200 Kb)
3) ligate into vectors
4) Transform E.coli cells
method of shearing DNA into pieces for insertion
sonication (vibrations)
genomic DNA library (def.)
collection of the total genomic DNA from a single organism stored in different vectors
sonication produces
equal, uniform fragments
RE isn’t used for shearing DNA because ___
DNA fragments would be different lengths
cDNA is ___
complementary DNA (from mRNA)
purpose of making cDNA is to __
clone all mRNAs from an organism
Making cDNA procedure
1)isolate mRNAS with poly A tails
2)reverse transpiratase to DNA from RNA
3)degrade mrna
4) dna poly copies ssdna to dsdna –> cDNA
5)ligate into vectors
6) transform cells
cDNA library (def.)
collection of the total genomic mRNA from a single organism stored in different vectors
cdna of mrna that had introns in DNA
cDNA would contain only the exons
in making cDNA libary, poly A tails are used to ___
isolate mrna
cDNA library would only contain genes that ____
were active and made mRNA
After BAC cloning/cDNA cloning, ____
DNA is sequenced before being stored at -70C (indexing the library-we know which DNA is in which clone)
BAC clones are used for ____
FISH
cDNAs clones are used for _____
GMOs
