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3. Nano LC ESI ion trap tandem MS ¼ºñ½º ÀýÂ÷ ¹× ¼ö°¡
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- 1D SDS-Page band ¹× 2D-Spot »óÅÂÀÇ ´Ü¹éÁú
In-gel digestion ÀÇ·Ú ½Ã
- Band ¹× gelÀ» 25~50 mM NH4HCO3 ¿¡ ÃæºÐÈ÷ ´ã¾Æ ¿À¼¼¿ä.
- LC-MS/MS ´ã´çÀÚ°¡ in-gel digestionÀ» ÇÏ°Ô µË´Ï´Ù. (1Àϼҿä)
- In-gel digestion µÈ peptide
-10ul ÀÌÇÏÀÇ 25~50 mM NH4HCO3 ¿¡ ´ã¾Æ¿À¼¼¿ä.
- Solution »óÅÂÀÇ protein
Solution »óÅ·Π³ÃÀå º¸°üÇÏ¿© °¡Á®¿À¼¼¿ä.
Sample »óÅ¿¡ µû¶ó in-solution ȤÀº in-gel solutionÀ¸·Î digestionÇÏ°Ô µË´Ï´Ù.
- ´Ü¹éÁúÀÇ ¼ö¿¡ µû¶ó LC-MS/MS running timeÀÌ ´Þ¶óÁý´Ï´Ù.
- 1~5°³ »çÀÌ ¾à 40~80 min, 5~10 »çÀÌ ¾à 80~120 min 10°³ ÀÌ»ó 120 min ÀÌ»ó
- ÀÇ·ÚÀÚ¿Í »óÀÇ ÈÄ °áÁ¤
- µ¥ÀÌÅÍ´Â MS/MS °á°ú¿¡ ÀÇÇØ ¾ò¾îÁø peptide·Î °Ë»öµÈ proteinÀÇ List, sequence, ºÐÀÚ·®, pI°ª, species,
peptide match ¼ö, scoreµîÀ¸·Î º¸¿©Áý´Ï´Ù. °¢°¢ÀÇ ³»¿ªÀ» excel ÆÄÀÏ·Î º¸³»µå¸³´Ï´Ù.
- ÁÖÀÇ »çÇ×
´ÙÀ½°ú °°Àº °æ¿ì ÁÁÀº MSµ¥ÀÌÅ͸¦ ¾ò±â°¡ ¾î·Æ½À´Ï´Ù.
- ´Ù·®ÀÇ salt°¡ Æ÷ÇÔµÈ protein/peptide(Àü 󸮽à ÁÖÀÇ¿ä¸Á)
- ÃæºÐÇÑ digestionÀÌ µÇÁö ¾ÊÀº protein ( 1/20~1/50 ºñÀ²·Î ÷°¡ ¿ä¸Á)
- sample¿¡ particle(¸ÕÁö ¹× ±âŸ À̹°Áú)ÀÌ Æ÷ÇԵǾî ÀÖÀ» °æ¿ì(½Ã½ºÅÛ ÀÚü¿¡ ¼Õ»ó)
- ½Ã·á´ç ºÐ¼® °¡°ÝÀº 19¸¸¿ø ÀÔ´Ï´Ù. ¼ºñ½º ½ÅûÀ» ÇϽñâ Àü, ¹Ýµå½Ã ¿¬±¸¿ø (ÀÌÀº¿µ :02-2123-6625,
)°ú ÇØ´ç ¼ºñ½º¿¡ ´ëÇØ¼ ²À ¹®ÀÇÇϽŠÈÄ, ¼ºñ½º ½ÅûÀ» ÇϽñ⠹ٶø´Ï´Ù.
- »ùÇà º¸³»½Ç ÁÖ¼Ò : ¼¿ï ¼´ë¹®±¸ ½ÅÃ̵¿ ¿¬¼¼´ëÇб³ »êÇÐÇùµ¿°ü 421È£.
- ºÐ¼® ¼ºñ½º ¿ä±ÝÀº ¿¬¼¼´ëÇб³ °èÁ·ΠÀԱݽÃÄÑ ÁÖ½Ã¸é µË´Ï´Ù. ¿¬¼¼´ëÇб³ ¸íÀÇÀÇ °è»ê¼°¡ ¹ßºÎµÇ¸ç, ¼¼±Ý°è»ê¼´Â ¹ß±ÞÀÌ ¾ÈµÇ°í, Ä«µå °áÀçµµ ¾ÈµË´Ï´Ù. °áÀç°èÁ¾ȳ»´Â ¸ÞÀÏ·Î µå¸³´Ï´Ù.
- ºÐ¼®¼ºñ½º ¼Ò¿ä½Ã°£
| Nano-LC-MS/MS 5 working day (1~5 sample) |
| destaining |
2 ½Ã°£ (per 10 spots) |
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| In-gel digestion |
12 ½Ã°£ |
|
| Poros desalting, spotting |
2 ½Ã°£ |
|
| LC-MS |
7½Ã°£ |
|
| Database searching |
3 ½Ã°£ |
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