QPCR - Lin Lab: Difference between revisions

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==Before the run:==

#Click on ~~“Shortcut to Sybr plate”~~ icon.

#Spin down plate 1 min at 3000 RPM in an empty 96 well plate using the Lin lab centrifuge

#Click on ~~“Instrument” tab and change volume of rxn. to 20uL~~. With new primers, may want to run the dissociation step to see if one peak (or more) is generated by the primers. This step adds about an extra hour to the run time. Can eliminate this step for primers which have already been analyzed in this way.

#Press power button.

#Go to File menu and then Save As. Go up one folder to ABI folder then to the Results folder, then save in the Saltiel folder ~~(in my folder).~~

#Open tray and put plate into tray.

~~#Close~~ the ~~program (Shortcut to Sybr plate window).~~

#Click on “7300 System Software” icon.

~~#Reopen the program through the 7300 System Software icon. Open the new saved file and~~ the ~~7300 system should then initialize~~.

#Click on open then in the directory Templates select SYBR Plate or SYBR Plate (no dissociation). With new primers, may want to run the dissociation step to see if one peak (or more) is generated by the primers. This step adds about an extra hour to the run time. Can eliminate this step for primers which have already been analyzed in this way.

#Under the Instrument tab, hit “Start.” ~~Note:~~ It takes a few minutes for the run to begin after “Start” is hit. Sometimes the run time is incorrectly calculated by the software.

#Click on “Instrument” tab and check that reaction volume is 20uL.

#Go to File menu and then Save As. Go up one folder to ABI folder then to the Results folder, then save in the Saltiel folder with the date as the filename.

#Under the Instrument tab, hit “Start.”

#Wait until a estimated run time is shown. It takes a few minutes for the run to begin after “Start” is hit. Sometimes the run time is incorrectly calculated by the software.

==After the run:==

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{| border = "1"

|Stage 1 || Stage 2 || Stage 3 | ~~colspan = "4"~~| Dissociation

|Stage 1 || Stage 2 || Stage 3 || || Dissociation

|-

| Rep 1 || Rep 1 || Rep 40 | ~~colspan = "4"~~| Stage 4

| Rep 1 || Rep 1 || Rep 40 || || Stage 4

|-

| || || | ~~colspan = "4"~~| Rep 1

| || || || || Rep 1

|-

| 50C || 95C || 95C || 60C || 95C || 60C || 95C

@@ Line 1: / Line 1: @@
 ==Before the run:==
-#Click on “Shortcut to Sybr plate” icon.
+#Spin down plate 1 min at 3000 RPM in an empty 96 well plate using the Lin lab centrifuge
-#Click on “Instrument” tab and change volume of rxn. to 20uL. With new primers, may want to run the dissociation step to see if one peak (or more) is generated by the primers.  This step adds about an extra hour to the run time.  Can eliminate this step for primers which have already been analyzed in this way.
+#Press power button.
-#Go to File menu and then Save As.  Go up one folder to ABI folder then to the Results folder, then save in the Saltiel folder (in my folder).
+#Open tray and put plate into tray.
-#Close the program (Shortcut to Sybr plate window).
+#Click on “7300 System Software” icon.
-#Reopen the program through the 7300 System Software icon.  Open the new saved file and the 7300 system should then initialize.
+#Click on open then in the directory Templates select SYBR Plate or SYBR Plate (no dissociation).  With new primers, may want to run the dissociation step to see if one peak (or more) is generated by the primers.  This step adds about an extra hour to the run time.  Can eliminate this step for primers which have already been analyzed in this way.
-#Under the Instrument tab, hit “Start.”    Note:  It takes a few minutes for the run to begin after “Start” is hit.  Sometimes the run time is incorrectly calculated by the software.
+#Click on “Instrument” tab and check that reaction volume is 20uL.
+#Go to File menu and then Save As.  Go up one folder to ABI folder then to the Results folder, then save in the Saltiel folder with the date as the filename.
+#Under the Instrument tab, hit “Start.”
+#Wait until a estimated run time is shown.  It takes a few minutes for the run to begin after “Start” is hit.  Sometimes the run time is incorrectly calculated by the software.
 ==After the run:==
@@ Line 24: / Line 27: @@
 {| border = "1"
-|Stage 1 || Stage 2 || Stage 3 | colspan = "4"| Dissociation
+|Stage 1 || Stage 2 || Stage 3 || || Dissociation
 |-
-| Rep 1 || Rep 1 || Rep 40 | colspan = "4"| Stage 4
+| Rep 1 || Rep 1 || Rep 40 || || Stage 4
 |-
-| || || | colspan = "4"| Rep 1
+| || || || || Rep 1
 |-
 | 50C || 95C || 95C || 60C || 95C || 60C || 95C