MEASUREMENT OF NUCLEIC ACIDQUALITYANDQUANTITY Flashcards
the agents used most frequently for visualization of bands after electrophoresis are
fluorescent dyes and silver stain
this dyes are specifically associated with nucleic acid
silver stain and fluorescent dye
quality is also known as
purity
3 mains methods to identify sample quality and quantity:
electrophoresis
spectrophotometry
fluorometry
this method to identify sample quality is for the presence of the bands
electrophoresis
this method to identify sample quality is to check absorbance
spectrophotometry
this method to identify sample quality is for the absorbance that is based on light that is emitted
fluorometry
this fluorescent dye is responsible for attachment or intercalation of the base pairs of the nucleic acid
eithidiun bromide
this fluorescent dye is specifically for electrophoresis
sybr green
this fluorescent dye is specifically for fluorometry
hoeschst dye
the appearance depends on the DNA isolated
electrophoresis
a high molecular weight genomic DNA should collect as a
bright band and low mobility
a high quality preparation of RNA will yield into two distinct band of rRNA, what are they?
tRNA and rRNA
The integrity of this bands is an indication of the integrity of the other RNA species present in the same sample
which concept is this
electrophoresis
after agarose gel electrophoresis, we can see different types of bands
super coild
nick
relax
linear plasmids
molecular ladders
a compact type of band that will travel farther in the gel
super coiled
a single stranded breaks in the bands
nick
a double stranded breaks in a band
relax
this band migrate according to the size of plasmin
linear plasmids
this band determines the size of the band
molecular ladders
most commonly used in visualizing nucleic acid
fluorescent dyes
an intercalation in visualizing nucleic acid in electrophoresis
fluorescent dyes
is a fluorescent dye that is the most widely used dye in early DNA and RNA analysis
ethidium bromide
ethidium bromide is also known as
3,8 diamino 5 ethyl 6 phenylphenanthridinium bromide
under excitation with UV light at 300 nm, EtBr in DNA will emit visible light at
590 nm
after soaking or running in dye, the dna illuminated with IV light will appear as what color in the gel
orange
used to visualize the image that can be capture
gel documentation box
MINORGROOVE–BINDING DYES used what stains
SYBR green
SYBR green that is used for double stranded DNA
SYBR I
SYBR green that is used for single stranded DNA
SYBR green II
this SYBR is used for both DNA and RNA staining
SYBR gold
SYBR green -has sensitivity of 2ng/mL. however, it is not specific for RNA, but it will bind double stranded DNA
true or false
true
difference of SYBR 1 green to EtBr
it does not intercalate between bases; it sots in the minor groove of the double helix
in agarose gel electrophoresis, SYBR staining is how many times more sensitive than ethidium bromide
25 to 100 times more
this dye is a cyanine dye of propietary structure
SYBR gold stain