Yeast DNA Prep protocol - source code: Difference between revisions
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< | <code><pre> | ||
#include "BioCoder.h" | #include "BioCoder.h" | ||
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Container eppendorf1 = new_container(EPPENDORF); | Container eppendorf1 = new_container(EPPENDORF); | ||
Container eppendorf2 = new_container(EPPENDORF); | Container eppendorf2 = new_container(EPPENDORF); | ||
//1. grow up yeast culture to appropriate density (near saturation) | //1. grow up yeast culture to appropriate density (near saturation) | ||
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//3. resuspend pellet in 200 ul breaking buffer | //3. resuspend pellet in 200 ul breaking buffer | ||
next_step(); | next_step(); | ||
measure_fluid(break_buffer, vol(200, UL), eppendorf1); | |||
resuspend(eppendorf1); | resuspend(eppendorf1); | ||
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// * 200 ul (@200 mg) glass beads | // * 200 ul (@200 mg) glass beads | ||
next_step(); | next_step(); | ||
measure_fluid(phe_chloro_iaa, vol(200, UL), eppendorf1); | |||
measure_solid(glass_beads, 200, MG, eppendorf1); | |||
comment("Wear gloves for this step!"); | comment("Wear gloves for this step!"); | ||
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//7. transfer 350 ml aqueous (top) layer to fresh eppendorf. | //7. transfer 350 ml aqueous (top) layer to fresh eppendorf. | ||
next_step(); | next_step(); | ||
measure_fluid(te, vol(200, UL), eppendorf1); | |||
centrifuge_phases_top(eppendorf1, speed(SPEED_MAX, RPM), RT, time(5, MINS), vol(350, UL), eppendorf2); | |||
//8. add 1 ml 95% ethanol and mix well, let sit for 10 minutes | //8. add 1 ml 95% ethanol and mix well, let sit for 10 minutes | ||
next_step(); | next_step(); | ||
measure_fluid(ethanol, vol(1, ML), eppendorf2); | |||
vortex(eppendorf2); | vortex(eppendorf2); | ||
store_for(eppendorf2, RT, time(10, MINS)); | store_for(eppendorf2, RT, time(10, MINS)); | ||
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//10. resuspend pellet in 50 ul TE buffer or water. | //10. resuspend pellet in 50 ul TE buffer or water. | ||
next_step(); | next_step(); | ||
first_option(); | |||
measure_fluid(te, vol(50, UL), eppendorf2); | |||
next_option(); | next_option(); | ||
measure_fluid(water, vol(50, UL), eppendorf2); | |||
end_option(); | end_option(); | ||
resuspend(eppendorf2); | resuspend(eppendorf2); | ||
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end_protocol(); | end_protocol(); | ||
} | } | ||
</ | |||
</pre></code> |
Latest revision as of 22:39, 19 November 2009
#include "BioCoder.h"
void main()
{
start_protocol("Yeast DNA Prep");
Fluid yeast_cul = new_fluid("yeast culture grown up to appropriate density", "near saturation");
Fluid break_buffer = new_fluid("breaking buffer", "2% (v/v) Triton X-100, 1% (w/v) SDS, 100 mM NaCl, 10 mM Tris-Cl, pH 8.0, 1 mM EDTA, pH 8.0");
Fluid phe_chloro_iaa = new_fluid("phenol : chloroform : isoamyl alcohol", "25:24:1", ICE_COLD);
Fluid te = new_fluid("TE buffer", "10 mM Tris-Cl, pH 8.0, 1 mM EDTA, pH 8.0");
Fluid ethanol = new_fluid("95% ethanol", ICE_COLD);
Fluid water = new_fluid("water");
Solid glass_beads = new_solid("glass beads");
Container eppendorf1 = new_container(EPPENDORF);
Container eppendorf2 = new_container(EPPENDORF);
//1. grow up yeast culture to appropriate density (near saturation)
//2. spin 1.5 mls of culture for 1 min in microfuge and aspirate off supernatant
first_step();
measure_fluid(yeast_cul, vol(1.5, ML), eppendorf1);
centrifuge_pellet(eppendorf1, speed(SPEED_MAX, RPM), RT, time(1, MINS));
//3. resuspend pellet in 200 ul breaking buffer
next_step();
measure_fluid(break_buffer, vol(200, UL), eppendorf1);
resuspend(eppendorf1);
//4. wear gloves and add:
// * 200 ul phenol:choloroform:isoamyl alcohol (25:24:1)
// * 200 ul (@200 mg) glass beads
next_step();
measure_fluid(phe_chloro_iaa, vol(200, UL), eppendorf1);
measure_solid(glass_beads, 200, MG, eppendorf1);
comment("Wear gloves for this step!");
//5. close cap tightly and vortex for 2.5 min.
// * Be careful when vortexing; label can be dissolved by the phenol.
// * Hold cap tightly so it doesn't open or spill.
next_step();
vortex(eppendorf1, time(2.5, MINS));
comment("Be careful when vortexing; label can be dissolved by the phenol.");
comment("Hold cap tightly so it doesn't open or spill.");
//6. add 200 ul TE buffer and spin for 5 min, in microfuge
//7. transfer 350 ml aqueous (top) layer to fresh eppendorf.
next_step();
measure_fluid(te, vol(200, UL), eppendorf1);
centrifuge_phases_top(eppendorf1, speed(SPEED_MAX, RPM), RT, time(5, MINS), vol(350, UL), eppendorf2);
//8. add 1 ml 95% ethanol and mix well, let sit for 10 minutes
next_step();
measure_fluid(ethanol, vol(1, ML), eppendorf2);
vortex(eppendorf2);
store_for(eppendorf2, RT, time(10, MINS));
//9. spin for 2 min, take off supernatant, and let dry upside down 10 min.
next_step();
centrifuge_pellet(eppendorf2, speed(SPEED_MAX, RPM), RT, time(2, MINS));
invert_dry(eppendorf2, RT, time(10, MINS));
//10. resuspend pellet in 50 ul TE buffer or water.
next_step();
first_option();
measure_fluid(te, vol(50, UL), eppendorf2);
next_option();
measure_fluid(water, vol(50, UL), eppendorf2);
end_option();
resuspend(eppendorf2);
comment("You can now use 1-2 ul of this crude yeast plasmid DNA prep to transform E. coli.");
end_protocol();
}