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本帖最后由 细胞海洋 于 2010-9-19 20:18 编辑
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$ K) ]9 y* `, y$ @& m1 O1 l" e6 S$ ]PCR 应该是分子生物学最基本的实验手段，但并不是每个实验都有一帆风顺的，基于PCR的实验越来越多，SSCP，定点突变，
( l9 b8 x" e! @& n3 z o' T% s; S等 PCR进行基因沉默的制备。比较实用的基因手段。大家根据需要选择下载，希望有所帮忙。
# x$ r$ Z& ^1 A$ nPreface .............................................................................................................v0 h3 `4 n# w( |* S! @8 Z5 C
Contributors .....................................................................................................xi, `* O4 n j/ W u3 ?
PART I. PERFORMING AND OPTIMIZING PCR
9 {1 q3 a& F- u* d) |5 d# d2 \4 W1 Polymerase Chain Reaction: Basic Principles and Routine Practice6 ]" Y/ m. J& P, S9 u5 G
Lori A. Kolmodin and David E. Birch .................................................. 3
8 B: H6 S8 F7 \( s) q2 Computer Programs for PCR Primer Design and Analysis
* w5 |( g* ?# P1 XBing-Yuan Chen, Harry W. Janes, and Steve Chen ........................ 19, Y9 O- P8 `/ D" K: Z0 e8 D
3 Single-Step PCR Optimization5 b# F+ Y8 O! E( s1 J# x
Using Touchdown and Stepdown PCR Programming
6 R7 a1 }9 g7 W9 e( T3 }0 rKenneth H. Roux .................................................................................. 31
; m9 S, f# {. E4 XL PCR Amplification of Long Targets from Genomic DNA7 e: B9 n* | g
Lori A. Kolmodin .................................................................................. 37
5 J/ a4 O7 Y( t9 z# S5 `/ c# U5 Coupled One-Step Reverse Transcription and Polymerase Chain
+ f _% F* U* q2 x& K1 {; GReaction Procedure for Cloning Large cDNA Fragments2 I% |( m4 `0 T1 o9 ]7 K
Jyrki T. Aatsinki ................................................................................... 53
9 V: a9 H O- m, o: }/ n6 [6 Long Distance Reverse-Transcription PCR u1 V4 x2 [" T
Volker Thiel, Jens Herold, and Stuart G. Siddell ............................. 593 [" X I" P" V
7 Increasing PCR Sensitivity for Amplification
* n6 \5 ]1 i7 _. m6 ?+ Y; R2 Efrom Paraffin-Embedded Tissues$ g& s/ i, e- r+ `* h% d1 F
Abebe Akalu and Juergen K. V. Reichardt ....................................... 67
2 f, G1 x! l& f8 GC-Rich Template Amplification by Inverse PCR:7 v% Y/ `2 K+ ~4 _: E' f
DNA Polymerase and Solvent Effects
- l7 F$ M2 H8 v' _9 {! r7 x8 \) cAlain Moreau, Da Shen Wang, Steve Forget, Colette Duez,5 }/ N% E- D, Y1 K; N& ?) z% T
and Jean Dusart............................................................................... 75# @ k1 y2 z3 K
9 PCR Procedure for the Isolation of Trinucleotide Repeats
/ B4 S Q' P& |2 q XTeruaki Tozaki ...................................................................................... 81# Q1 n$ F. g3 Q3 Q( H2 Y- O
10 Methylation-Specific PCR2 k: B. D5 w0 I; U$ z* q8 _
Haruhiko Ohashi .................................................................................. 910 w" n& D U& V
11 Direct Cloning of Full-Length Cell Differentially Expressed Genes
1 ~' i+ G }- Gby Multiple Rounds of Subtractive Hybridization
; y- ~/ y! \; r% f6 V! c! ?Based on Long-Distance PCR and Magnetic Beads
* @6 o0 ~$ C# l( K- m5 IXin Huang, Zhenglong Yuan, and Xuetao Cao ................................ 99
) U3 f% v) u( r8 E8 WPART II. CLONING PCR PRODUCTS. w- P5 }& S) \& k( F& T) M9 B
12 Cloning PCR Products: An Overview
) w7 D5 \7 R+ Q/ l9 e5 U, mBaotai Guo and Yuping Bi ................................................................ 111
3 A( _) d/ H3 x/ j' h13 Using T4 DNA Polymerase to Generate Clonable PCR Products* p( ~3 ?6 ~% q" o8 v7 n4 S
Kai Wang ............................................................................................. 121
) {. U/ }( ^4 B) l$ E* o14 Enzyme-Free Cloning of PCR Products' b0 v* o0 {& ^2 {4 Y+ X' r
and Fusion Protein Expression" c* a# w3 w0 h. p
Brett A. Neilan and Daniel Tillett ..................................................... 125
# ^8 W% P( C6 n0 b- ?15 Directional Restriction Site-Free Insertion of PCR Products! X! ?5 i9 A1 g. E" Z
into Vectors
) v0 W' t% z- E# e I. h9 NGuo Jun Chen .................................................................................... 133
$ o6 v- x) J# E16 Autosticky PCR:
( w) ?$ q# J6 i2 K" @/ Q6 VDirectional Cloning of PCR Products with Preformed 5' Overhangs3 n" n. G/ G6 m% Z& U1 ~. G
József Gál and Miklós Kálmán......................................................... 141
8 J; a5 h1 q8 D) @. E17 A Rapid and Simple Procedure for Direct Cloning
2 E7 t; N! V8 y0 c0 y0 f+ `of PCR Products into Baculoviruses
4 N; l* ^' H3 _6 z% j: L& DTamara S. Gritsun, Michael V. Mikhailov,/ _; D( A* |; Y/ A" y
and Ernest A. Gould ...................................................................... 153
" Y! o% `# y6 e9 b6 |: r- OPART III. MUTAGENESIS AND RECOMBINATION
}0 Y8 }! U- x$ o% k2 I! N7 E18 PCR Approaches to DNA Mutagenesis and Recombination:
2 N- t D# s8 M! G& J' zAn Overview" H: z- G+ \' N1 q! A# V: ?
Binzhang Shen ................................................................................... 167( f3 i, g8 j9 P b2 [4 j4 D
19 In-Frame Cloning of Synthetic Genes Using PCR Inserts2 r! l9 C& a& v( x
James C. Pierce ................................................................................. 175
' f) p4 \ R( M. h; \% s; N20 Megaprimer PCR
4 s+ | w1 L9 }8 P& ySailen Barik ........................................................................................ 1898 T9 F* r) c2 g
21 PCR-Mediated Recombination:0 l; s: m% n% G
A General Method Applied to Construct Chimeric Infectious
9 t- _+ n/ E x9 L/ {9 jMolecular Clones) v: _( k$ U$ Y" J1 x3 U' O, r9 Y0 |4 i
Guowei Fang, Barbara Weiser, Aloise Visosky, Timothy Moran,
( u: z) V2 a6 C/ P( j9 eand Harold Burger ......................................................................... 1972 v* n- T) P$ Z1 |. x
22 PCR Method for Generating Multiple Mutations at Adjacent Sites
H$ y4 X- M) d. R+ g- |Jiri Adamec ......................................................................................... 2076 }$ v' `# ^3 N) T- k
23 A Fast Polymerase Chain Reaction-Mediated Strategy for Introducing
) V- }+ G6 @9 w' S1 S' p% v- WRepeat Expansions into CAG-Repeat Containing Genes8 Q W* f8 T. i
Franco Laccone ................................................................................. 2176 G( w" S* E) I3 F, ~* _
24 PCR Screening in Signature-Tagged Mutagenesis of Essential Genes. w3 |9 q. Z( X: K( d
Dario E. Lehoux and Roger C. Levesque ....................................... 225- W% O, @7 q. \) r/ Y
25 Staggered Extension Process (StEP) In Vitro Recombination
: d" n h7 K. N; n0 L6 `0 G% Z6 bAnna Marie Aguinaldo and Frances Arnold ................................... 235
0 t4 }% G8 O+ w6 n5 O' Q26 Random Mutagenesis by Whole-Plasmid PCR Amplification! f* y/ t$ q' p+ [, l) ~7 J
Donghak Kim and F. Peter Guengerich .......................................... 241
! U4 N2 v/ W6 |! r1 r" `7 VPART IV. CLONING UNKNOWN NEIGHBORING DNA. ?! X0 S+ {3 Y5 A0 O! b
27 PCR-Based Strategies to Clone Unknown DNA Regions K+ N L- c" r1 G' p- S
from Known Foreign Integrants: An Overview3 j3 V" }# g |
Eric Ka-Wai Hui, Po-Ching Wang, and Szecheng J. Lo ................ 2490 {8 [" m- K& e
28 Long Distance Vectorette PCR (LDV PCR)( v$ X) o& o9 B8 _
James A. L. Fenton, Guy Pratt, and Gareth J. Morgan ................. 275
8 X$ o$ @9 N5 f$ x' q29 Nonspecific, Nested Suppression PCR Method
8 x6 R" A& ?# ^4 P$ w, lfor Isolation of Unknown Flanking DNA (“Cold-Start Method”)9 ?$ w0 I/ T. G/ N4 a U
Michael Lardelli .................................................................................. 2856 X# J5 ^! r2 _5 d
30 Inverse PCR: cDNA Cloning2 e3 z& j& z4 f3 Y- Y6 R4 M
Sheng-He Huang ................................................................................ 293
x: q1 w6 |# E) \( c+ S31 Inverse PCR: Genomic DNA Cloning$ w5 [) {' |* F7 x. q# H
Ambrose Y. Jong, Anna T’ang, De-Pei Liu,
5 z9 ^$ s# V3 q4 y, ^' E" sand Sheng-He Huang .................................................................... 301+ \! m9 L! ?1 D8 U0 W) ?
32 Gene Cloning and Expression Profiling by Rapid Amplification. B" O ~: M* P3 ~0 q2 l# y
of Gene Inserts with Universal Vector Primers
5 ?' L' Z; E" nSheng-He Huang, Hua-Yang Wu, and Ambrose Y. Jong .............. 3097 N( I' H: p3 F; u) ?! Q
33 The Isolation of DNA Sequences Flanking Tn5 Transposon Insertions7 f. L& Q& Q% X1 ~7 | q7 O
by Inverse PCR
; A! r# _: t9 V9 a: P( U2 nVincent J. J. Martin and William W. Mohn ...................................... 315
/ E, h& d6 ?9 \7 e3 i2 \& I2 l1 c) c34 Rapid Amplification of Genomic DNA Sequences Tagged
" H. [* h* m+ `6 z" iby Insertional Mutagenesis
M' b2 y1 G! H: ZMartina Celerin and Kristin T. Chun ................................................ 325
" i( R) |' {. _* o2 `& e35 Isolation of Large Terminal Sequences of BAC Inserts Based
. n6 N- H" D2 Eon Double-Restriction-Enzyme Digestion Followed
5 `) n3 J/ ?( A# _by Anchored PCR0 p" A9 f: t0 R
Zhong-Nan Yang and T. Erik Mirkov ............................................... 337# d; R9 z9 c1 |5 b
36 A “Step Down” PCR-Based Technique for Walking
# w/ r N( ^: o: V3 P! V2 {Into and the Subsequent Direct Sequence Analysis
/ ^. s; D4 t$ V3 F( z* gof Flanking Genomic DNA
: _( y( O6 a8 S1 N6 C( S( _Ziguo Zhang and Sarah Jane Gurr .................................................. 343! d/ |" I' v7 |" s+ h& g f9 `
PART V. LIBRARY CONSTRUCTION AND SCREENING* \& n2 ~3 v7 ]2 I+ g5 \; }2 C4 i
37 Use of PCR in Library Screening: An Overview
6 K! D' ~1 E5 Y% C+ f; O5 yJinbao Zhu .......................................................................................... 353% ^! D9 [5 x8 c+ t
38 Cloning of Homologous Genes by Gene-Capture PCR% h3 Y3 U5 S9 F3 ~+ A3 e6 F5 i
Renato Mastrangeli and Silvia Donini ............................................. 359
* r0 Y2 p9 f8 M; V+ h J& u9 P39 Rapid and Nonradioactive Screening of Recombinant Libraries by PCR
8 C' `( L, m2 K2 a0 T( OMichael W. King ................................................................................. 377* Q8 }: r2 I: w L
40 Rapid cDNA Cloning by PCR Screening (RC-PCR) m# l! i8 S7 i P, Z2 s
Toru Takumi ....................................................................................... 3852 _% @4 L3 m" V5 n8 M& `
41 Generation and PCR Screening of Bacteriophage λ Sublibraries. Z4 w3 |7 c1 h8 d
Enriched for Rare Clones (the “Sublibrary Method”); P H3 f( R% ^; w/ a$ J
Michael Lardelli .................................................................................. 391, Y) e# q2 z8 P# @5 s( }1 |8 {0 ~6 e
42 PCR-Based Screening for Bacterial Artificial Chromosome Libraries
2 q$ z+ \, @+ `. f. nYuji Yasukochi ................................................................................... 4012 `$ C* @1 @( ^- D: p
43 A 384-Well Microtiter-Plate-Based Template Preparation
% U2 }' s' W, |; r% m2 ] Qand Sequencing Method
, M. b! N) b% f+ N6 }Lei He and Kai Wang ......................................................................... 411
; x W8 ]! R3 ~, k+ I44 A Microtiter-Plate-Based High Throughput PCR Product
$ B% x4 m6 r/ G' _! C* c aPurification Method. ?/ B6 M( P7 v3 ~( Q. P
Ryan Smith and Kai Wang ................................................................ 4175 \* \) @! X+ y' V' v
Index ............................................................................................................ 423* Q, Y8 \! N2 F- I% m% _% C8 z* g
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发表于 2010-9-19 18:24
