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̙33V0 B(  T3f    s *[`p@ T [<4___PPT9 T CEA WV(WmS|~ NvzLv $\vQ/fv~LvTÀLv-Nؚh Vdk(W$v~ހQShKm0RvQl U_,g (Wck8^bNSOQg\h 0 X8 xD8 x$C$T$(  S H  0޽h ? ̙33PW0 ( \ t  p  Hi/|2NA?@:  0[P  ck8^bNSOQCEAWVh 2 (   H  0޽h ? ̙33X0 bZ( 0h R  s *s`sp  Hz &KA?v  0`[P   ÀLv$v~~-NCEAWVvh(2 (   H  0޽h ? ̙33Y0 ,(     s *[ T [<4___PPT9 > (W$vSu@Slye YhT@ -N g\ϑ$v~ހkYu0(uupOe0 yr_vb/ghKm0RُN$v~ހ -NCEAWVvlU_,g /fSs $veglyvsQ.0< J $ZC$(>   H  0޽h ? ̙33ZZ0  (  L p  Hl}MA?`P  0[  "ÀLv`YhT@-NCEAWVvh(2 (    H  0޽h ? ̙33 [0 B: D( tH0h0 Dl D C (u   P D c (A 2 D s `v0e0e@R     D 0xzP DMDWVY>fP[:1YvhKm00(2 CA   H D 0޽h ? 3fff \0 T(  Tl T C ؄`   l T C   H T 0޽h ? 3fff]0 F>(  f  >e0#4Add-pcr0  6[ `Z ^ ]_>f:yRT-PCR(Differential display RT-PCR).0/GC  $ H  0޽h ? 3fff80___PPT10.=@Qqa0 @d( |@(@ dl d C    l d C p `    H d 0޽h ? 3fff b0 P`(  `l ` C d`   l ` C (Pb   H ` 0޽h ? 3fff6 0  6(   ~  s *|s `   x  c $(`   H  0޽h ? 3fff80___PPT10.uH0 $ (   :  3 Anp :  3 A]` `w   0  `  "Bcl I RFLP@SuA  2 8     H  0޽h ? 380___PPT10.eD0 ,(  ,R , C *A$RFLP-11B , s *޽h ? 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SouthernpSBgN"(2G    H 4 0޽h ? 3fff___PPT10i.bЏ.+D='  = @B +R 0  R(   ~  s *:`     s **`  "p`PpH  0޽h ? 3fff80___PPT10.0<0  |(   D  C A3 0H  0޽h ? 3fff6 0  6(   ~  s *8V`   x  c $ 0   H  0޽h ? 3fff80___PPT10.0&'B0   (   J  C "A! DGGEH  0޽h ? 3fff6 0 0 6(   x  c $     ~  s *`   H  0޽h ? 3fff80___PPT10.U<0 ^V@ (   R  C *A"heterodp\  C 4A&transgenomicpH  0޽h ? 380___PPT10.ec< 0 P<(  ~  s * `   ~  s *$    H  0޽h ? 3fff80___PPT10.@a60 `6(    C bA'H180px-Melting_Curve_Analysis_Graphs` 0l  C DA(*MeltingCurveAnalysis R H  0޽h ? 380___PPT10.e9V0 0 p0(  x  c $     x  c $  P2   H  0޽h ? 3fff80___PPT10.F}P0  (   X   0AI?H   0޽h ? 3fff|0 ,$(  X  0AJ?   0K p  PCRKm^o:yHr4 0(2  ^B @ 6D@P^B  6DP`  0Pl 0 P P  S1 0(2   0Y 0 P  S2 0(2 H  0޽h ? 3fffy0 ( v l  C B   l  C F `    H  0޽h ? 3fff$ z0  $(   r   S PPP    r   S 0Qp   H   0޽h ? 3fff80___PPT10.90  0(  ^ S     # l^f^f ?   Most researchers are well versed in the performance of PCR. However, I usually emphasize that the performance of Real-Time PCR requires an emphasis on each step. STEPS 1) Prepare the Master Mixture and add all components to the reaction tube. 2) Denature the Template to generate single-stranded DNA. Usually performed at 95 oC for 30 to 60 s. 3) Annealing temperature. The temperature is lowered in order to promote the binding of the primers to its complimentary target. Generally ranges between 50 and 65 oC. l P G FI     H  0޽h ? ̙330 rj (  ^ S    d # l^f^f ?   Starts with a Picture of the Annealed Template. 4) Extension Step - Between 65 and 72 oC. In many cases, this step is linked to the annealing temperature. When done this way, it is considered a 2 step PCR. Lead into next slide It is repeated to generate an exponential growth. H4z5*W   H  0޽h ? ̙330 WO (  ^ S    I # lp!^f^f ?   kThis demonstrates the power of exponential growth. 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