NYMU Taipei/phptest

From 2007.igem.org

(Difference between revisions)
 
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Line 4: Line 4:
<script language="javascript">
<script language="javascript">
-
function CheckId(SI,SV){
+
function CheckId(SI,SV,CI,CV,VI,L,R){
var SI = SI;
var SI = SI;
var SV = SV;
var SV = SV;
 +
var CI = CI;
 +
var CV = CV;
 +
var VI = VI;
 +
var L  = L;
 +
var R  = R;
 +
var DVV = 3/CV;
 +
var MI = R * (SI/SV);
//document.write("insert is v1, and vector is v2");
//document.write("insert is v1, and vector is v2");
-
alert("insert size = " + SI + "(bp)\n");
+
alert("insert size = " + SI + "bp\n" + "vector size = " + SV + "bp\n" + "vector concentration = " + CV + "ug/uL\n" + "digested vector volume = " + DVV + "uL\n" + "insert mass = " + MI + "ug");
}
}
Line 17: Line 24:
<form name=form1>
<form name=form1>
-
     <INPUT type="text" name="SI" > size (bp) of insert part (SI)<BR>
+
     <INPUT type="text" name="SI"> size (bp) of insert part (SI)<BR>
-
     <INPUT type="text" name="SV" > size (bp) of vector part (SV)<BR>
+
     <INPUT type="text" name="SV"> size (bp) of vector part (SV)<BR>
-
     <INPUT type="text" name="CI" > concentration (ug/uL) of insert (CI)<BR>
+
     <INPUT type="text" name="CI"> concentration (ug/uL) of insert (CI)<BR>
-
     <INPUT type="text" name="CV" > concentration (ug/uL) of vector (CV)<BR>
+
     <INPUT type="text" name="CV"> concentration (ug/uL) of vector (CV)<BR>
-
     <INPUT type="text" name="VI" > volume (uL) of insert for gel separation (VI)<BR>
+
     <INPUT type="text" name="VI"> volume (uL) of insert for gel separation (VI)<BR>
-
     <INPUT type="text" name="L" > volume (uL) of insert for ligation (L)<BR>
+
     <INPUT type="text" name="L"> volume (uL) of insert for ligation (L)<BR>
-
     <input type=button value="送出" Onclick="CheckId(SI.value, SV.value, CI.value, CV.value, VI.value, L.value);">
+
    <INPUT type="text" name="R">  ligation ratio<BR>
 +
     <input type=button value="送出" Onclick="CheckId(SI.value, SV.value, CI.value, CV.value, VI.value, L.value, R.value);">
</form>
</form>
</body>
</body>
Line 33: Line 41:
** BSA (2uL)
** BSA (2uL)
** plasmid + ddH2O (14uL = VI + water, 1 <= VI <= 14)
** plasmid + ddH2O (14uL = VI + water, 1 <= VI <= 14)
-
** (2) 300 ng = CV (ug/uL) * VV (uL) * 1000, VV = 0.3/CV
+
** for vector
 +
*** we assume 300 ng for digestion and 90% loss
 +
*** thus, amplify 10X from 300 ng to 3,000 ng = 3 mg
 +
*** 3,000 ng = CV (ug/uL) * 1000 (ng/ug) * DVV, DVV (digested vector volume) = 3/CV
----
----
* total gel separation volume is 50 uL
* total gel separation volume is 50 uL
----
----
* total ligation volume is 10uL
* total ligation volume is 10uL
-
** vector weight is 300 ng
+
** (MI/MV) = R * (SI/SV)
 +
** assume mass of vector (MV) is 300 ng
 +
*** MI = R * (SI/SV) * MV = 300R * (SI/SV) (ng)
** 10X buffer (1uL)
** 10X buffer (1uL)
** ligase (1uL)
** ligase (1uL)
Line 44: Line 57:
----
----
* formula
* formula
-
** (1) 300 * SI * 3 * 10 (損失因子) / SV = CI * VI * L /50
+
** '''300''' * SI * 3 (ligation ratio) * 10 (損失因子) / SV = CI * VI * L /50
-
 
+
** mass of vector (MV) is '''300''' ng
 +
** SI (bp) is size of insert
 +
** SV (bp) is size of vector
 +
** VI (uL) is volume of insert
** VV (uL) is volume of vector
** VV (uL) is volume of vector
 +
** CI (ng/uL) is concentration of insert
** CV (ng/uL) is concentration of vector
** CV (ng/uL) is concentration of vector
 +
** R is ranged from 1 to 10

Latest revision as of 10:07, 16 September 2007

size (bp) of insert part (SI)
size (bp) of vector part (SV)
concentration (ug/uL) of insert (CI)
concentration (ug/uL) of vector (CV)
volume (uL) of insert for gel separation (VI)
volume (uL) of insert for ligation (L)
ligation ratio

  • total digestion volume is 20 uL (typical)
    • enzyme 1 and enzyme 2 (1uL * 2)
    • 10X buffer (2uL)
    • BSA (2uL)
    • plasmid + ddH2O (14uL = VI + water, 1 <= VI <= 14)
    • for vector
      • we assume 300 ng for digestion and 90% loss
      • thus, amplify 10X from 300 ng to 3,000 ng = 3 mg
      • 3,000 ng = CV (ug/uL) * 1000 (ng/ug) * DVV, DVV (digested vector volume) = 3/CV
  • total gel separation volume is 50 uL
  • total ligation volume is 10uL
    • (MI/MV) = R * (SI/SV)
    • assume mass of vector (MV) is 300 ng
      • MI = R * (SI/SV) * MV = 300R * (SI/SV) (ng)
    • 10X buffer (1uL)
    • ligase (1uL)
    • VV + L + water (8uL)
  • formula
    • 300 * SI * 3 (ligation ratio) * 10 (損失因子) / SV = CI * VI * L /50
    • mass of vector (MV) is 300 ng
    • SI (bp) is size of insert
    • SV (bp) is size of vector
    • VI (uL) is volume of insert
    • VV (uL) is volume of vector
    • CI (ng/uL) is concentration of insert
    • CV (ng/uL) is concentration of vector
    • R is ranged from 1 to 10
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