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PDF电子书：RNA Purification
目录：
- w9 r4 V& a3 N# {Selecting a Purification Strategy . . . . . . . . . . . . . . . .. . . . . . . . . 1985 R% a% ] m6 a, `' i
Do Your Experiments Require Total RNA or mRNA? . . . . . 1982 ]" o4 L6 C( y/ M
Is It Possible to Predict the Total RNA Yield from: D+ j3 ?! \0 K
a Certain Mass of Tissue or Number of Cells? . . . . . . . . 201
, M3 Q* Q) t- `" @0 G/ R( }Is There Protein in Your RNA Preparation, and
6 o( T* Z, |6 y( tIf So, Should You Be Concerned? . . . . . . . . . . . . . . . . . . . . 202
6 P3 c9 Y* s- z- WIs Your RNA Physically Intact? Does It Matter? . . . . . . . . . . 202
! c% L' J( q0 }4 o5 n TWhich Total RNA Isolation Technique Is Most
) y) K! H0 D+ S! X5 q8 SAppropriate for Your Research? . . . . . . . . . . . . . . . . . . . . . 203$ ~! `3 b. s7 C7 H2 z& [7 v
What Protocol Modifications Should Be Used for
+ j" u& k7 }, C! b7 |$ C! tRNA Isolation from Difficult Tissues? . . . . . . . . . . . . . . . . 207
& ]! |5 [2 L) H. K7 K+ `( h" }' hIs a One-Step or Two-Step mRNA-(poly(A) RNA)-
' a1 e. r$ q+ G2 ?; d; {4 Z8 pPurification Strategy Most Appropriate for Your
9 _9 h; G+ ~! y% w' V. S7 H, uSituation? . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2099 F u$ {- O0 \- i- Y: ^) g
How Many Rounds of Oligo(dT)–Cellulose
$ i* b" }% U) w* ~Purification Are Required? . . . . . . . . . . . . . . . . . . . . . . . . . 210
: V, T* d f* I9 x% GWhich Oligo(dT)–Cellulose Format Is Most: |: p3 I5 u- x$ }4 ?
Appropriate? . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 210; Y$ K6 Z& e1 }+ e' R! B
Can Oligo(dT)–Cellulose Be Regenerated and Reused? . . . 211
% @( b- O% f2 [. o$ z2 s2 n' eCan a Kit Designed to Isolate mRNA Directly from- v, \( N: d5 \' F/ |' L1 N
the Biological Sample Purify mRNA from Total RNA? . . . 212) A% ?; e& F- |/ t" O7 g
Maximizing the Yield and Quality of an RNA Preparation . . . 2121 ^; V! U& I3 `0 s4 K: ~( A
What Constitutes “RNase-Free Technique”? . . . . . . . . . . . . 2128 e, j# o, g' g% ~3 W: }
How Does DEPC Inhibit RNase? . . . . . . . . . . . . . . . . . . . . . . 213! W, i! O4 d8 B: C3 j0 ^& W
How Are DEPC-Treated Solutions Prepared? Is
' \2 L. J% u4 e6 F" M- v6 ZMore DEPC Better? . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 213
3 {% F: X# I. \8 ~' RShould You Prepare Reagents with DEPC-Treated Water,& t$ \- e2 A* f/ F0 Q
or Should You Treat Your Pre-made Reagents with
" t4 |0 F6 X3 ]- z% z6 ^9 m, h- `DEPC? . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 214: O/ w, G C2 @1 U6 D6 o9 z5 H
How Do You Minimize RNA Degradation during Sample
, u0 M$ O3 C$ eCollection and Storage? . . . . . . . . . . . . . . . . . . . . . . . . . . . . 214: l9 b7 [. t Y# S
How Do You Minimize RNA Degradation during Sample" d8 Z) m% X* V# ~
Disruption? . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 215- Q% U2 e1 L! n6 q i4 f& v
Is There a Safe Place to Pause during an RNA/ E1 {3 |/ q: c
Purification Procedure? . . . . . . . . . . . . . . . . . . . . . . . . . . . . 218
! E' l3 d, H9 n& x( H" X9 iWhat Are the Options to Quantitate Dilute RNA
0 e1 e3 d" y; P, FSolutions? . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 218
! h$ K; ^* ^" J* B8 cWhat Are the Options for Storage of Purified RNA? . . . . . . . 219
) m8 `8 J. O# P# eTroubleshooting . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2208 f8 R! B* |9 T* o h
A Pellet of Precipitation RNA Is Not Seen at the End of! H# m# Z& ]2 w) J. W" n* Q' K
the RNA Purification . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 220
& V" e, B8 {% K( j) }A Pellet Was Generated, but the Spectrophotometer4 ~# h, W3 J# a* X" U" s0 D
Reported a Lower Reading Than Expected, or Zero% ]3 y; I( K5 a( S' l
Absorbance . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 221
5 L. Q7 f% n( X, W0 L# h9 y' |RNA Was Prepared in Large Quantity, but it Failed. F0 i& m. n- E4 f% ~
in a Downstream Reaction: RT PCR is an
U* t X6 q, S. W5 v' ^Example . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 221) J. u# t* T i4 i; A" |; ?, q- `
My Total RNA Appeared as a Smear in an Ethidum
; j; I) a$ M! P* m& d( fBromide-stained Denaturing Agarose Gel; 18S and" V8 @6 C* \3 |- _) t* j2 U4 a
28S RNA Bands Were not Observed . . . . . . . . . . . . . . . . 2220 g" E4 r& v- R1 L# `; r
Only a Fraction of the Original RNA Stored at -70°C+ H( S; M3 h8 E9 R3 i6 F3 N9 H' B
Remained after Storage for Six Months . . . . . . . . . . . . . . 222
( d$ r: O9 t, J+ M- GBibliography . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 222 |
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发表于 2011-10-25 19:26
发表于 2011-10-26 08:59
