NYMU Taipei/phptest
From 2007.igem.org
(Difference between revisions)
| Line 22: | Line 22: | ||
<form name=form1> | <form name=form1> | ||
| - | <INPUT type="text" name="SI" > | + | <INPUT type="text" name="SI"> size (bp) of insert part (SI)<BR> |
| - | <INPUT type="text" name="SV" > | + | <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="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);"> | <input type=button value="送出" Onclick="CheckId(SI.value, SV.value, CI.value, CV.value, VI.value, L.value);"> | ||
</form> | </form> | ||
| Line 46: | Line 47: | ||
---- | ---- | ||
* total ligation volume is 10uL | * total ligation volume is 10uL | ||
| - | ** vector | + | ** (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) | ||
Revision as of 10:00, 16 September 2007
- 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 * 10 (損失因子) / SV = CI * VI * L /50
- vector weight 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
