- ºü¸£°í ½¬¿î ½ÇÇè°úÁ¤ - 10 ºÐ ¹ÝÀÀÀ̸é ÃæºÐ
- »ý¹°ÇÐÀû È°¼ºÀ» À¯ÁöÇÒ¼ö ÀÖ´Â nondenaturing extraction
- ¸ðµç IMAC Resin¿¡ ÀûÇÕÇÏ¸ç ºü¸¥ his-tagged proteinÀÇ Á¤Á¦¸¦ ½ÇÇö
- ¹ü¿ëÀû- ´Ù¾çÇÑ scaleÀÇ ´Ü¹éÁú ÃßÃâ°ú ´Ù¾çÇÑ tag (his-, GST-, FLAG-, HA-tagged protein°ú ÇÔ²² »ç¿ë)¿¡ »ç¿ë °¡´É
- Bacterial, yeast, mammalian, ¶Ç´Â baculovirus-infected cells·ÎºÎÅÍ protein ÃßÃâ
- SonicationÀ̳ª ´Ù¸¥ cell lysis bufferº¸´Ù ´õ È¿°úÀû
´Ù¾çÇÑ ºÐÀÚ·®ÀÇ affinity-tagged proteinÀ» Á¤Á¦Çϱâ À§ÇÏ¿© mammalian cell pellet extraction »Ó¸¸ ¾Æ´Ï¶ó
bacteria¿¡µµ TALON xTractor Buffer¸¦ »ç¿ë °¡´ÉÇÏ´Ù. ¶ÇÇÑ TALON xTractor Buffer Kit¿¡´Â lysozyme°ú DNase°¡
Æ÷ÇԵǾî ÀÖ¾î bacteria ¼¼Æ÷º®°ú ¸·À» ¿ÏÀüÈ÷ Æı«ÇÒ ¼ö Àֱ⠶§¹®¿¡ ³ôÀº ºÐÀÚ·®ÀÇ ´Ü¹éÁúÀ»
È¿À²ÀûÀ¸·Î ÃßÃâ °¡´É ÇÏ´Ù. º» Á¦Ç°Àº inclusion body¿¡¼ ºÒ¿ëÇؼº ´Ü¹éÁúÀ» ÃßÃâÇϱâ À§ÇØ »ç¿ëÇÒ ¼ö ÀÖ´Ù.
Protein Extraction with xTractor Buffer than Sonication
Extraction
Method |
Fraction |
Protein
(mg) |
Activity |
Results |
Sonication |
Total lysate |
28.6 |
872 |
Sonication provides lower yield & activity |
Purified protein |
0.6 |
2,115 |
xTractor Buffer Kit |
Total lysate |
122.1 |
2,277 |
xTractor provides higher yield & activity |
Purified protein |
0.9 |
3,054 |
For this comparison of protein extraction methods, LacZ protein was extracted from a 1.7 g pellet of E. coli cells expressing his-tagged protein.
Lysates were purified on a TALON column and eluted with 150 nM imidazole. Protein activity was determined by measuring luminescence units.