XIE/3:2026/Jan: Difference between revisions
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*Purification :The incubated samples were mixed with 400 μL of phosphate‑buffered saline (PBS) and transferred into 50 kDa molecular‑weight‑cut‑off (MWCO) centrifugal ultrafiltration units. Samples were centrifuged at 10,000 × g for 3 min, the retentate was brought back to volume with PBS, and the centrifugal ultrafiltration step was repeated; this wash/centrifugation cycle was performed a total of three times | *Purification :The incubated samples were mixed with 400 μL of phosphate‑buffered saline (PBS) and transferred into 50 kDa molecular‑weight‑cut‑off (MWCO) centrifugal ultrafiltration units. Samples were centrifuged at 10,000 × g for 3 min, the retentate was brought back to volume with PBS, and the centrifugal ultrafiltration step was repeated; this wash/centrifugation cycle was performed a total of three times | ||
===琼脂糖凝胶电泳=== | ===琼脂糖凝胶电泳(1.23)=== | ||
#2.4g琼脂糖加入到120 ml（0.5X TBE）中，微波炉加入至完全溶解；用 0.5X TBE 补足，再加入 1.2ml 1M MgCl2 和 12 ul Goldview；倒入模具，插上梳子。 | #2.4g琼脂糖加入到120 ml（0.5X TBE）中，微波炉加入至完全溶解；用 0.5X TBE 补足，再加入 1.2ml 1M MgCl2 和 12 ul Goldview；倒入模具，插上梳子。 | ||
#取Scaffold、Origami（50nM）、Origami（100nM）、Origami（150nM）各 10 ul， 各加入 Loudingbuffer 2 ul，涡旋混匀。 | #取Scaffold、Origami（50nM）、Origami（100nM）、Origami（150nM）各 10 ul， 各加入 Loudingbuffer 2 ul，涡旋混匀。 | ||
#Scaffold、Origami（50nM）、Origami（100nM）、Origami（150nM）依次上样，90V 3h（电泳液：0.5X TBE + 10mM MgCl2） | #Scaffold、Origami（50nM）、Origami（100nM）、Origami（150nM）依次上样，90V 3h（电泳液：0.5X TBE + 10mM MgCl2） | ||
[[File:T_Origami_2026-01-23.jpg|thumb|center|400px]] | [[File:T_Origami_2026-01-23.jpg|thumb|center|400px]] | ||
结论：150 nM 浓度太低 | <span style="color:red;">结论：150 nM 浓度太低</span> | ||
<div style="color: black !important; font-family: 'Times New Roman', SimSun, serif !important, line-height:1.5;"> | |||
==1.25== | |||
*<big><span style="color:red;">'''Origami folding'''</span></big> | |||
===折叠三种浓度的Origami（100nM,190nM,300nM）=== | |||
*300 nM 的Origami是通过超滤浓缩获得 | |||
{| class="wikitable" | |||
|+ 190 nM | |||
! Samples | |||
! Volume | |||
|- | |||
| Scaffolld (10uM) | |||
| 1.9 ul | |||
|- | |||
| Staples (10uM) | |||
| 9.5 ul X 9 | |||
|- | |||
| PBS (10X) | |||
| 10 ul | |||
|- | |||
| Water | |||
| 2.6 ul | |||
|} | |||
{| class="wikitable" | |||
|+ 100 nM X 4 | |||
! Samples | |||
! Volume | |||
|- | |||
| Scaffolld (1uM) | |||
| 10 ul | |||
|- | |||
| Staples (10uM) | |||
| 5 ul X 9 | |||
|- | |||
| PBS (10X) | |||
| 10 ul | |||
|- | |||
| Water | |||
| 35 ul | |||
|} | |||
*Program folding in PCR thermocycler : The mixed sample was put on a thermal ramp starting with '''a rapid heat denaturation at 80 °C for 5 min followed by cooling from 80 °C to 60 °C over 20 min, then slow cooling from 60 °C to 24 °C over 14 h'''. | |||
Latest revision as of 08:18, 26 January 2026
1.22[edit]
- Origami folding
折叠三种浓度的Origami（50nM,100nM,150nM）[edit]
| Samples | Volume |
|---|---|
| Scaffolld (1uM) | 5 ul |
| Staples (10uM) | 2.5 ul X 9 |
| PBS (10X) | 10 ul |
| Water | 62.5 ul |
| Samples | Volume |
|---|---|
| Scaffolld (1uM) | 10 ul |
| Staples (10uM) | 5 ul X 9 |
| PBS (10X) | 10 ul |
| Water | 35 ul |
| Samples | Volume |
|---|---|
| Scaffolld (1uM) | 15 ul |
| Staples (10uM) | 7.5 ul X 9 |
| PBS (10X) | 10 ul |
| Water | 7.5 ul |
- Program folding in PCR thermocycler : The mixed sample was put on a thermal ramp starting with a rapid heat denaturation at 80 °C for 5 min followed by cooling from 80 °C to 60 °C over 20 min, then slow cooling from 60 °C to 24 °C over 14 h.
- Purification :The incubated samples were mixed with 400 μL of phosphate‑buffered saline (PBS) and transferred into 50 kDa molecular‑weight‑cut‑off (MWCO) centrifugal ultrafiltration units. Samples were centrifuged at 10,000 × g for 3 min, the retentate was brought back to volume with PBS, and the centrifugal ultrafiltration step was repeated; this wash/centrifugation cycle was performed a total of three times
琼脂糖凝胶电泳(1.23)[edit]
- 2.4g琼脂糖加入到120 ml（0.5X TBE）中，微波炉加入至完全溶解；用 0.5X TBE 补足，再加入 1.2ml 1M MgCl2 和 12 ul Goldview；倒入模具，插上梳子。
- 取Scaffold、Origami（50nM）、Origami（100nM）、Origami（150nM）各 10 ul， 各加入 Loudingbuffer 2 ul，涡旋混匀。
- Scaffold、Origami（50nM）、Origami（100nM）、Origami（150nM）依次上样，90V 3h（电泳液：0.5X TBE + 10mM MgCl2）
结论：150 nM 浓度太低
1.25[edit]
- Origami folding
折叠三种浓度的Origami（100nM,190nM,300nM）[edit]
- 300 nM 的Origami是通过超滤浓缩获得
| Samples | Volume |
|---|---|
| Scaffolld (10uM) | 1.9 ul |
| Staples (10uM) | 9.5 ul X 9 |
| PBS (10X) | 10 ul |
| Water | 2.6 ul |
| Samples | Volume |
|---|---|
| Scaffolld (1uM) | 10 ul |
| Staples (10uM) | 5 ul X 9 |
| PBS (10X) | 10 ul |
| Water | 35 ul |
- Program folding in PCR thermocycler : The mixed sample was put on a thermal ramp starting with a rapid heat denaturation at 80 °C for 5 min followed by cooling from 80 °C to 60 °C over 20 min, then slow cooling from 60 °C to 24 °C over 14 h.