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PDF电子书：RNA Purification
目录：" F9 |. j8 L* ?- C+ |; z2 D0 o
Selecting a Purification Strategy . . . . . . . . . . . . . . . .. . . . . . . . . 198
7 T* t, r7 \* I& b6 [Do Your Experiments Require Total RNA or mRNA? . . . . . 198
% w& D$ {% o* F. d) x1 c) |5 kIs It Possible to Predict the Total RNA Yield from
0 e! h0 B% h7 E- Q% M; ga Certain Mass of Tissue or Number of Cells? . . . . . . . . 201& B% _7 b7 x+ h" u; X2 \; h& _
Is There Protein in Your RNA Preparation, and
+ ^! n6 l0 ^* hIf So, Should You Be Concerned? . . . . . . . . . . . . . . . . . . . . 202
9 n- _( V6 n5 r5 `0 W3 eIs Your RNA Physically Intact? Does It Matter? . . . . . . . . . . 202# b9 l8 B1 I4 s' z9 a7 W
Which Total RNA Isolation Technique Is Most
4 q# q9 g B) k6 a. c4 N% i' YAppropriate for Your Research? . . . . . . . . . . . . . . . . . . . . . 203" |: v$ ^6 j( ^; T$ x8 E5 T. J
What Protocol Modifications Should Be Used for
6 Q3 d" d* O/ z) [. B5 M1 \+ `- ]7 T+ MRNA Isolation from Difficult Tissues? . . . . . . . . . . . . . . . . 2071 u8 X5 G3 k7 [2 V6 n! q `
Is a One-Step or Two-Step mRNA-(poly(A) RNA)-% X6 A! h2 v8 |& Z( y( X6 a
Purification Strategy Most Appropriate for Your# | T% x3 H2 S7 r. U, G1 a6 Z
Situation? . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2094 [+ S+ p4 K9 w* ?- ]
How Many Rounds of Oligo(dT)–Cellulose
! S' @" {* ]5 ? OPurification Are Required? . . . . . . . . . . . . . . . . . . . . . . . . . 2107 k6 W% g3 |- l
Which Oligo(dT)–Cellulose Format Is Most1 Q% Y# C8 ^) u3 t" I, K* C
Appropriate? . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 210
# j* J: `) x' f& ICan Oligo(dT)–Cellulose Be Regenerated and Reused? . . . 211
7 H7 l7 W# f) p ], ]Can a Kit Designed to Isolate mRNA Directly from
# q; \8 ] P, G* Wthe Biological Sample Purify mRNA from Total RNA? . . . 212
9 j2 {6 `7 Z& R" A F: X4 uMaximizing the Yield and Quality of an RNA Preparation . . . 212; k+ y9 J9 p5 T* y, {4 E$ R; E
What Constitutes “RNase-Free Technique”? . . . . . . . . . . . . 2129 N3 ^) T/ S) O9 G4 o. z
How Does DEPC Inhibit RNase? . . . . . . . . . . . . . . . . . . . . . . 213
9 t. e8 |! [" G) \7 b) P1 vHow Are DEPC-Treated Solutions Prepared? Is
5 K: I2 r1 v. O9 mMore DEPC Better? . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 213
& S" o J; U/ F" r& RShould You Prepare Reagents with DEPC-Treated Water,
8 o5 F3 K& N6 ^ zor Should You Treat Your Pre-made Reagents with9 }2 C. C) T3 u% y: s
DEPC? . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2148 H" p$ I' e4 M) u: f
How Do You Minimize RNA Degradation during Sample
# l# ~# J s6 q% ~9 U, w; ^Collection and Storage? . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2143 @: M% j' J3 |. Q3 a8 j
How Do You Minimize RNA Degradation during Sample5 `! q: W6 t+ Y9 Z" j1 y5 _
Disruption? . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 215/ ~# c% ]4 D. A6 B+ P/ c; B. y8 A: o
Is There a Safe Place to Pause during an RNA1 Y2 f6 h, H" Z
Purification Procedure? . . . . . . . . . . . . . . . . . . . . . . . . . . . . 218# C& S' ~ R) O
What Are the Options to Quantitate Dilute RNA, h3 t1 Y" Z9 I% ]9 t
Solutions? . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 218
# G* O6 U! S( @ g e* j2 QWhat Are the Options for Storage of Purified RNA? . . . . . . . 2192 F Y. ^" q) r% ^ `
Troubleshooting . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 220
4 W- f: N4 P! G; ?A Pellet of Precipitation RNA Is Not Seen at the End of
9 z; v w& s& X$ s2 bthe RNA Purification . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 220
& c# Z @6 o# N1 `4 W. }A Pellet Was Generated, but the Spectrophotometer
% P1 ?; N% L" Y+ G; ^Reported a Lower Reading Than Expected, or Zero0 T$ V3 a' ^ H3 X7 V
Absorbance . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 221. s L5 B) c9 C, d# n
RNA Was Prepared in Large Quantity, but it Failed9 H4 N- R$ S( Z1 r0 r
in a Downstream Reaction: RT PCR is an: \( w }" `1 }; a2 |" @) |$ p' w
Example . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 221
1 U+ w* e( _ \! ^My Total RNA Appeared as a Smear in an Ethidum
1 h" _# H! W$ p: d- i' ? \Bromide-stained Denaturing Agarose Gel; 18S and
" K; r. X& I0 ?7 M4 i8 H' C* [% C, i28S RNA Bands Were not Observed . . . . . . . . . . . . . . . . 222
% M$ v1 T; m5 u3 vOnly a Fraction of the Original RNA Stored at -70°C# _0 m" c4 ]2 y* l4 x9 W; \2 M
Remained after Storage for Six Months . . . . . . . . . . . . . . 222+ l# I5 w7 h A
Bibliography . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 222 |
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发表于 2011-10-25 19:26
发表于 2011-10-26 08:59
