Differences

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--- oxfordjuly072008:start [2014/05/15 17:25] – magiero
+++ oxfordjuly072008:start [2015/01/30 21:19] (current) – magiero
@@ Line 10: / Line 10: @@
 ==== - Molecular Adaptor ====
+•[0141] …molecular adaptor…facilitates the interaction between the pore and the nucleotides.  The adaptor typically alters the charge of the barrel or channel of the pore or specifically interacts with or binds to nucleotides \\
+•[0142] The adaptor preferably constricts the barrel or channel so that it may interact with the nucleotides. \\
+•[0143] The adaptor is typically cyclic.  The adaptor preferably has the same symmetry as the pore.
 ==== - Covalent Attachment ====
+• [0149] The adaptor is covalently attached to the pore.\\
+• [0150] The site of covalent attachment is selected such that the adaptor is positioned at or near residue 139 of SEQ ID NO:2. **(This denotes the vertical position of the adaptor)**\\
 ==== - Positioning of the Adaptor ====
@@ Line 29: / Line 36: @@
 ==== - Apparatus ====
-• [0197] An example apparatus consists of two chambers separated by the pore carrying membrane.
+• [0197] An example apparatus consists of two chambers separated by the pore carrying membrane.\\
 • [0199] Voltage clamp is preferred to patch clamp. **What is the reason for this preference?**
@@ Line 54: / Line 61: @@
 ==== - Exonuclease ====
-• [0212] Attach an exonuclease to the pore.  An exonuclease is a sequence snipping enzyme is processes DNA like a ribbon sequentially chopping off (**digesting**) nucleotides one after another.\\
+• [0212] Attach an exonuclease to the pore.  An exonuclease is a sequence snipping enzyme. It processes DNA like a ribbon sequentially chopping off (**digesting**) nucleotides one after another.\\
 • [0215] Preferable to covalently attach the exonuclease to the pore.\\
 • [0216] Preferable exonuclease rates are 1, 10, 100, 500, 1000 nucleotides per second.\\
@@ Line 84: / Line 91: @@
 === - Data Analysis and Acquisition ===
-• [0236] Axopatch 200B patch clamp amplifier used.  4-pole 10-kHz Bessel filter.  Sampled at 20 kHz by PC equipped with Digidata 1440A A/D (Axon instruments) running CampEx 10 software (Molecular Devices).\\
+• [0236] Axopatch 200B patch clamp amplifier used.  4-pole 10-kHz Bessel filter.  Sampled at 20 kHz by PC equipped with Digidata 1440A A/D (Axon instruments) running ClampEx 10 software (Molecular Devices).\\
 • [0237] Event histograms are constructed as follows…\\
 • [0238] Run statistics on a moving window consisting of WT points.\\
@@ Line 103: / Line 110: @@
 === - High Positions of Attachment--Residues 115 to 121 ===
-• [0261] In the 115 position in <m>\beta</m>-barrel baseline was too noise and showed large current fluctuations.  Maybe due to movement of cyclodextrin in barrel.\\
+• [0261] In the 115 position in the <m>\beta</m>-barrel the baseline was too noisy and showed large current fluctuations.  Maybe due to movement of cyclodextrin in barrel.\\
 • [0263] Further down in 119 position baseline got cleaner, but still showed some spikes.\\
@@ Line 121: / Line 128: @@
 | dCMP    | 44 pA            | Smallest current block      |
-From Fig. 15 baseline current is around 67 pA.\\
+From Fig. 15 **baseline current** is around 67 pA.\\
 • [0280] As you increase the applied voltage you get increase in passing currents and not necessarily equal shifts in the histograms (Fig. 18).\\
 • [0281] Histogram overlab (at different voltages) looks pretty small (Fig. 19).\\
-• [0282] The dTMP histogram looks the sharpest.  Why?
+• [0282] The dTMP histogram looks the sharpest.  Why?\\
-• [0283] Because its **dwell time** is the largest.  The longer (in time) the signal it generates the more samples we accrue and hence the more accurate our sampling of it (therefore the sharper the histogram). A summary of avg. dwell times in Fig. 20 at 130 mV and 800 mM KCl
+• [0283] Because its **dwell time** is the largest.  The longer (in time) the signal it generates the more samples we accrue and hence the more accurate our sampling of it (therefore the sharper the histogram). A summary of avg. dwell times in Fig. 20 at 130 mV and 800 mM KCl (just eyeballed the max. dwell times from Fig. 20).
+| Species | Avg Dwell Time |  Max Dwell Time |
+| dGMP    |  5.0 ms        |  25 ms          |
+| dTMP    | 19.8 ms        |  80 ms          |
+| dAMP    |  5.6 ms        |  25 ms          |
+| dCMP    |  7.3 ms        |  48 ms          |
-| Species | Dwell Time |
+**Recall**, sampling time is 20-kHz so every 0.05 ms.  Avg. dwell time is 9.4 ms, so on average they are **oversampling** by 188 times.\\
-| dGMP    |  5.0 ms    |
-| dTMP    | 19.8 ms    |
-| dAMP    |  5.6 ms    |
-| dCMP    |  7.3 ms    |
 === - Low Salt Operating Conditions ===