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¹«¼¼Æ÷ ´Ü¹éÁú ÇÕ¼º°è

Human Cell-Free Protein Expression System

-

Á¦Á¶»ç Á¦Ç°ÄÚµå Á¦Ç°¸í ¿ë·® °¡°Ý
(ºÎ°¡¼¼º°µµ)
ºñ°í »ç¿ëÀڸŴº¾ó
Takara
3281
Human Cell-Free Protein Expression System
°ü·ÃÇмú ±¸¸ÅÇϱâ
10 ȸ
531,200¿ø 
664,000¿ø
°¡°ÝÇÒÀÎ
09.02 ~ 10.31
3281_e.v1807Da.pdf
Takara
3290
pT7-IRES His-N DNA
°ü·ÃÇмú ±¸¸ÅÇϱâ
20 ¥ìg
276,000¿ø 
345,000¿ø
°¡°ÝÇÒÀÎ
09.02 ~ 10.31
3290_DS.v2109Da.pdf
Takara
3291
pT7-IRES His-C DNA
°ü·ÃÇмú ±¸¸ÅÇϱâ
20 ¥ìg
276,000¿ø 
345,000¿ø
°¡°ÝÇÒÀÎ
09.02 ~ 10.31
3291_DS.v2109Da.pdf
Takara
3292
pT7-IRES Myc-N DNA
°ü·ÃÇмú ±¸¸ÅÇϱâ
20 ¥ìg
276,000¿ø 
345,000¿ø
°¡°ÝÇÒÀÎ
09.02 ~ 10.31
3292_DS.v2109Da.pdf

  • »ç¶÷ ¼¼Æ÷ÁÖ À¯·¡ÀÇ ¼¼Æ÷ ÃßÃâ¾×À» ÀÌ¿ëÇÑ ¹«¼¼Æ÷ ´Ü¹éÁúÇÕ¼º½Ã½ºÅÛ
  • IRES¼­¿­ÀÇ ÀÌ¿ë, ¹ø¿ªÁõ°­ÀÎÀÚ È¿°ú¸¦ ÀÌ¿ëÇØ ÇÕ¼º·® Çâ»ó
  • 150 kDaÀÌ»óÀÇ °íºÐÀÚ ´Ü¹éÁú ÇÕ¼º¿¡µµ ÀÀ¿ë °¡´É
  • º°µµ ÆǸŵǴ vector¸¦ ÀÌ¿ëÇÏ¿© His-Tag³ª c-Myc-Tag¸¦ Ãß°¡ÇÑ ¸ñÀû ´Ü¹éÁú ¹ßÇöµµ °¡´É
Á¦Ç°¼³¸í
Human Cell-Free Protein Expression System(code 3281)Àº »ç¶÷ ¼¼Æ÷ÁÖ À¯·¡ÀÇ ¼¼Æ÷ ÃßÃâ¾×À» ÀÌ¿ëÇÑ ¹«¼¼Æ÷ ´Ü¹éÁú ÇÕ¼º½Ã½ºÅÛÀ̸ç, ÇϳªÀÇ Æ©ºê¿¡¼­ RNA Àü»ç·ÎºÎÅÍ ´Ü¹éÁú ÇÕ¼º±îÁö ½Ç½ÃÇÒ ¼ö ÀÖ´Ù.
º» ½Ã½ºÅÛÀÇ Cell Lysate¿¡´Â in vitro¿¡¼­ ´Ü¹éÁú ÇÕ¼º¹ÝÀÀ¿¡ ÇÊ¿äÇÑ °¢Á¾ ÀÎÀÚ(¸®º¸¼Ø, ¹ø¿ª °³½Ã • ½ÅÀå ÀÎÀÚ, tRNAµî)°¡ Æ÷ÇԵǾî ÀÖ´Ù. ÀÌ cell lysate¿¡ ¸ñÀûÀ¯ÀüÀÚ¸¦ Ŭ·Î´×ÇÑ T7-IRES Vector¿Í T7 RNA Polymerase, Mixture-3(APTµî°ú ¾Æ¹Ì³ë»ê·ù), Mixture-2(¹ø¿ªÁõ°­ÀÎÀÚ)¸¦ ÷°¡ÇÏ´Â °£´ÜÇÑ °úÁ¤¸¸À¸·Î ½ÇÇèÀÌ °¡´ÉÇÏ´Ù(±×¸² 1). º» ½Ã½ºÅÛ¿¡¼­´Â ¸ñÀû À¯ÀüÀÚÀÇ Àü»ç ¹ÝÀÀ°ú ¹ø¿ª ¹ÝÀÀÀÌ µ¿ÀÏ Æ©ºê³»¿¡¼­ ÀÌ·ç¾îÁö¹Ç·Î, 2´Ü°è ¹ÝÀÀ°úÁ¤¿¡¼­ ¿ì·ÁµÇ´Â mRNAÀÇ ºÐÇظ¦ ÃÖ¼ÒÈ­ ÇÒ ¼ö ÀÖ´Ù. ¶Ç, ÀϹÝÀûÀÎ Åä³¢ ¸Á»óÀûÇ÷±¸(rabbit reticulocyte)¸¦ ÀÌ¿ëÇÑ ¹«¼¼Æ÷ ´Ü¹éÁú ÇÕ¼º½Ã½ºÅÛº¸´Ù ÇÕ¼º È¿À²ÀÌ ³ô°í, ÇÕ¼º·®ÀÇ Áõ°¡¸¦ ±â´ëÇÒ ¼ö ÀÖ¾î(±×¸² 2), 150~200 kDaÀÇ °íºÐÀÚ ´Ü¹éÁú ÇÕ¼º¿¡µµ ÀÌ¿ëÇÒ ¼ö ÀÖ´Ù(±×¸² 3).
ÀÌ ½Ã½ºÅÛÀº Single-Step ¹ÝÀÀÀ» ÀÌ¿ëÇÑ High-throughput ´Ü¹éÁú ÇÕ¼º ¿Ü¿¡µµ ¼¼Æ÷ ¹ßÇö½Ã ¼÷ÁÖ ¼¼Æ÷ÀÇ ¼ºÀå ¹× ¼¼Æ÷ ±â´É¿¡ ¿µÇâÀ» ¹ÌÄ¡´Â µ¶¼º ´Ü¹éÁú ÇÕ¼º¿¡µµ ÀÀ¿ëÇÒ ¼ö ÀÖ´Ù. ±×¸² 1
±×¸²1. Human Cell-Free Expression System¸ð½Äµµ

¹ÝÀÀ¾×£ÛCell Lysates*, T7 RNA Polymerase, Mixture-1, Mixture-2(¹ø¿ª Áõ°­ ÀÎÀÚ), Mixture-3(ATPµîÀ̳ª ¾Æ¹Ì³ë»ê·ù)£Ý¿¡ ¸ñÀû À¯ÀüÀÚ¸¦ Ŭ·Î´×ÇÑ pT7-IRES Vector¸¦ ÷°¡ÇÏ´Â °£ÆíÇÑ 1 Æ©ºê ¹ÝÀÀÀ¸·Î, mRNAÀÇ Àü»ç·ÎºÎÅÍ ´Ü¹éÁú ÇÕ¼º±îÁö¸¦ one-stepÀ¸·Î ½Ç½ÃÇÒ ¼ö ÀÖ´Ù.
* Cell Lysate ¿¡´Âin vitro ´Ü¹éÁú ÇÕ¼º ¹ÝÀÀ¿¡ ÇÊ¿äÇÑ ¸®º¸¼Ø, ¹ø¿ª °³½Ã • ½ÅÀå ÀÎÀÚ, tRNA µîÀÇ °¢Á¾ ÀÎÀÚ°¡ Æ÷ÇԵǾî ÀÖ´Ù.

º» ½Ã½ºÅÛÀ¸·Î ÀÌ¿ëÇϱâ À§Çؼ­ His-Tag³ªc-Myc-Tag¸¦ Æ÷ÇÔÇÑ ¹ßÇöº¤Å͵µ º°µµ ÆǸÅÇÏ°í ÀÖ´Ù. ÀÌ·¯ÇÑ ¹ßÇö º¤ÅÍ´Â T7ÇÁ·Î¸ðÅÍ¿Í EMCV IRES¼­¿­ ¸»´Ü¿¡ His-Tag³ªc-Myc-TagµîÀÇ Tag ¼­¿­, Factor XaÀý´Ü ¼­¿­, MCS, polyA ½Ã±×³Î, T7 terminator¸¦ Æ÷ÇÔÇÏ°í ÀÖ´Ù. Tag Á¾·ù, ¼­¿­ À§Ä¡°¡ ´Ù¸¥ 3Á¾·ùÀÇ º¤ÅÍ°¡ ÀÖÀ¸¸ç, ¸ñÀû¿¡ ¸Â´Â ÃÖÀûÀÇ º¤Å͸¦ ¼±ÅÃÇÒ ¼ö ÀÖ´Ù.
¿¹¸¦ µé¸é, pT7-IRES His-N DNA(code 3290)´Â ¸ñÀû ´Ü¹éÁúÀÇN¸»´Ü¿¡, pT7-IRES His-C DNA(code 3291)´Â ¸ñÀû ´Ü¹éÁúÀÇ C¸»´Ü¿¡, His-Tag¸¦ °¡Áö°í ÀÖ´Â ´Ü¹éÁú ¹ßÇöÀÌ °¡´ÉÇÏ´Ù. His-Tag¸¦ ÀÌ¿ëÇÏ¿© Á¤Á¦ ÈÄ, ¹ßÇöµÈ ´Ü¹éÁú·ÎºÎÅÍHis-Tag¸¦ Á¦°ÅÇÒ ¼ö ÀÖ´Â Factor XaÀý´Ü ¼­¿­µµ Æ÷ÇÔÇÏ°í ÀÖ´Ù. ¶Ç, pT7-IRES Myc-N DNA(code 3292)´Â ¸ñÀû ´Ü¹éÁúÀÇN¸»´Ü¿¡c-Myc-Tag¸¦ °¡Áö°í ÀÖ´Â ´Ü¹éÁúÀÇ ¹ßÇöÀÌ °¡´ÉÇÏ°í, µ¿ÀÏÇÑFactor XaÀý´Ü ¼­¿­À» °¡Áö°í ÀÖ´Ù. (Vector map ÂüÁ¶)
ÀÌ·¯ÇÑ º¤ÅÍÀÇ MCS¿¡ ÇÁ·¹ÀÓÀ» ¸ÂÃß¾î »ðÀÔÇÑ ¸ñÀû À¯ÀüÀÚ´ÂT7ÇÁ·Î¸ðÅÍÀÇ Á¦¾îÇÏ¿¡¼­ EMCV IRES¸¦ °¡Áö´Â RNA·Î Àü»çµÇ¾î, EMCV IRESÀÇ ¹ø¿ª °³½Ã ÃËÁø È¿°ú¿¡ ÀÇÇØ º» cell-free protein expression(¹«¼¼Æ÷ ´Ü¹éÁú ¹ßÇö)½Ã½ºÅÛ¿¡¼­ È¿À²ÀûÀÎ °í¹ßÇöÀÌ °¡´ÉÇÏ´Ù.
³»¿ë
Human Cell-Free Protein Expression System(code 3281)
1. Cell Lysate*1 100 ¥ìl
2. Mixture-1 60 ¥ìl
3. Mixture-2*2 10 ¥ìl
4. Mixture-3*2 20 ¥ìl
5. T7 RNA Polymerase (200 U/¥ìl) 10 ¥ìl
6. pT7-IRES Vector (0.5 ¥ìg/¥ìl) 20 ¥ìl
7. Control Vector*3 (0.3 ¥ìg/¥ìl) 5 ¥ìl

*1 : »ç¿ë Á÷Àü¿¡ ³ì¿©¼­ ÇÇÆêµîÀ¸·Î ºÎµå·´°Ô ÃæºÐÈ÷ ¼¯Àº ÈÄ Áï½Ã »ç¿ëÇÏ°í, »ç¿ë ÈÄ¿¡´Â Áï½Ã -80¡É¿¡ º¸°üÇÑ´Ù.
(ÁÖÀÇ) 5ȸ Á¤µµÀÇ µ¿°áÀ¶Çطδ ¼º´ÉÀÌ ÀúÇϵÇÁö ¾Ê´Â °ÍÀ» È®ÀÎÇÏ¿´Áö¸¸, °¡´ÉÇÑ ºÐÁÖÇÏ¿© º¸°üÇÒ °ÍÀ» ±ÇÀåÇÕ´Ï´Ù..
*2 : Mixture-2¹×Mixture-3´Â ´Ü¹éÁúÀ» Æ÷ÇÔÇÏ°í ÀÖÀ¸¹Ç·Î, °úµµÇÑ ±³¹Ý µî ´Ü¹éÁúÀÇ ½ÇÈ°À» ÃÊ·¡ÇÏ´Â Á¶ÀÛÀº ÇÇÇØ¾ß ÇÑ´Ù. Mixture-2¿¡ Æ÷ÇԵǾî ÀÖ´Â ´Ü¹éÁú¿¡´ÂHN-Tag ¼­¿­ÀÌ Ã·°¡µÇ¾î ÀÖ´Ù.
*3 : ¥â-Galactosidase À¯ÀüÀÚ°¡ Æ÷ÇԵǾî ÀÖ´Ù.
º¸Á¸
code 3281 ; -80¡É
code 3290,3291,3292 ; -20¡É
Vector Map
±×¸² 2
* Human Cell-Free Protein Expression System¿¡ Æ÷ÇÔ µÇ¾î ÀÖÀ½.
±×¸² 3
Vector ¼­¿­Á¤º¸
pT7-IRES(FastaÇü½Ä)
pT7-IRES_His-N(FastaÇü½Ä)
pT7-IRES_His-C(FastaÇü½Ä)
pT7-IRES_Myc-N(FastaÇü½Ä)

FastaÇü½Ä £º »óµ¿¼­¿­°Ë»öÀÇ µ¥ÀÌŸº£À̽º³ª ¼­¿­À» Ãë±ÞÇÏ´Â °¢Á¾ ÅøÀ» »ç¿ëÇÒ ¶§¿¡ »ç¿ëµÇ´Â ÆÄÀÏ Çü½ÄÀ¸·Î, ù¹ø° ÇàÀÇ >µÚ¿¡ ÆÄÀÏ Á¤º¸(º¤Å͸í)¸¦ ³Ö´Â´Ù. ¼­¿­Àº 60¿°±â¸¶´Ù ÇàÀ» ¹Ù²Û´Ù.
IRES ¼­¿­ ÀÌ¿ë°ú µ¶ÀÚÀûÀÎ ¹ø¿ª Áõ°­ ÀÎÀÚ¸¦ ÀÌ¿ëÇÑ ³ôÀº ¹ø¿ª È¿À² À¯Áö
pT7-IRES Vector·ÎºÎÅÍ Àü»çµÇ´Â ¸ñÀû À¯ÀüÀÚRNA¿¡´Â ´Ü¹éÁúÀÇ ¹ø¿ª °³½Ã¸¦ ÃËÁøÇÏ´Â IRES¼­¿­ÀÌ Æ÷ÇԵǾî ÀÖ´Ù. ¶ÇÇÑ, ¹ÝÀÀ¾×¿¡ ÷°¡µÇ´Â °íÀ¯ÀÇ ¹ø¿ª Áõ°­ ÀÎÀÚ´Â, ´Ü¹éÁú ÇÕ¼º°ú ÇÔ²² ºÒÈ°¼ºÈ­µÈ ¹ø¿ª °³½Ã ÀÎÀÚ¸¦ ÀçÈ°¼ºÈ­ÇÏ¿©, ¹ø¿ª ÀúÇϸ¦ ¾ïÁ¦ÇÏ¿© ³ôÀº ¹ø¿ª È¿À²À» À¯ÁöÇÑ´Ù. ÀÌ·¯ÇÑ ¹ø¿ªÈ¿À² Áõ°­È¿°ú¿¡ ÀÇÇØ, º» ½Ã½ºÅÛ¿¡¼­´Â ±âÁ¸ÀÇ Åä³¢ ¸Á»óÀûÇ÷±¸(Rabbit reticulocyte)¸¦ ÀÌ¿ëÇÑ ¹«¼¼Æ÷ ´Ü¹éÁú ÇÕ¼º ½Ã½ºÅÛÀÇ ´ÜÁ¡À̾ú´ø ³·Àº ÇÕ¼º·üÀÌ Çâ»ó½ÃÄ×°í, 150 kDaÀÌ»óÀÇ °íºÐÀÚ ´Ü¹éÁúÀÇ ÇÕ¼º¿¡ ÀÀ¿ë °¡´ÉÇÔÀ» È®ÀÎÇÏ¿´´Ù. (±×¸²2, ±×¸²3).
±×¸² 4

±×¸²2. Human Cell-Free Protein Expression System°ú Åä³¢ ¸Á»óÀûÇ÷±¸¸¦ ÀÌ¿ëÇÑ ¹«¼¼Æ÷ ´Ü¹éÁú ÇÕ¼º ½Ã½ºÅÛ¿¡¼­ÀÇ ¥â-galactosidase ÇÕ¼º·® ºñ±³
°¢°¢ÀÇ ÇÁ·ÎÅäÄÝ¿¡ µû¶ó ¥â-galactosidase ÇÕ¼º ¹ÝÀÀÀ» È­ÇÐ ¹ß±¤ ±âÁúÀ» ÀÌ¿ëÇØ ÃøÁ¤Çß´Ù. ±× °á°ú, Åä³¢ ¸Á»óÀûÇ÷±¸¸¦ ÀÌ¿ëÇÑ ¹«¼¼Æ÷ ´Ü¹éÁú ÇÕ¼º¿¡ ºñÇØ, »ç¶÷ ¼¼Æ÷ÁÖ À¯·¡ÀÇ º» ½Ã½ºÅÛ¿¡¼­ ¸Å¿ì ³ôÀº ¼öÁØÀÇ ´Ü¹éÁúÀÌ ÇÕ¼ºµÇ´Â °ÍÀ» È®ÀÎÇÏ¿´´Ù.

±×¸² 5

±×¸²3. º» ½Ã½ºÅÛÀ» ÀÌ¿ëÇÑ °íºÐÀÚ ´Ü¹éÁú(170 kDa, 200 kDa) ¹ßÇö
º» ½Ã½ºÅÛÀ» ÀÌ¿ëÇØ °íºÐÀڴܹéÁúÀ» ÇÕ¼ºÇÏ¿´´Ù. SDS-PAGEÈÄ, CBB¿°»öÀ¸·Î ¸ñÀû ´Ü¹éÁúÀ» °ËÃâÇÑ °á°ú, ¿øÇÏ´Â Å©±â(170 kDa, 200 kDa)ÀÇ ´Ü¹éÁúÀ» È®ÀÎÇÒ ¼ö ÀÖ¾ú´Ù.
Lane 1: Negative Control
Lane 2: Human Dicer(200 kDa)
Lane 3: Human eIF4G(170 kDa)
¿ëµµ
  • °íºÐÀÚ ´Ü¹éÁú ÇÕ¼º
  • ±â´É¼º ´Ü¹éÁú ÇÕ¼º
  • µ¶¼º ´Ü¹éÁú ÇÕ¼º
  • cDNA»ê¹°ÀÇ Æ÷°ýÀûÀÎ ½ºÅ©¸®´×
  • ´Ü¹éÁú/´Ü¹éÁú »óÈ£ÀÛ¿ë ºÐ¼®
  • ´Ü¹éÁú/DNA»óÈ£ÀÛ¿ë Çؼ®
  • ´Ü¹éÁúÀÇ º¯ÀÌ µµÀÔ ºÐ¼®
ÁÖÀÇ »çÇ×
´ç»ç Ãë±Þ Á¦Ç°Àº ¸ðµÎ ¿¬±¸¿ëÀ¸·Î ÆǸÅÇÏ°í ÀÖ½À´Ï´Ù. »ç¶÷, µ¿¹° ´ëÇÑ ÀÇ·á, ÀÓ»ó Áø´Ü¿ëÀ¸·Î´Â »ç¿ëÇÏÁö ¾Ê°Ô ÁÖÀÇÇØ ÁֽʽÿÀ. ¶ÇÇÑ, ½ÄÇ°, È­ÀåÇ°, °¡Á¤¿ëÇ° µîÀ¸·Îµµ »ç¿ëÇÒ ¼ö ¾ø½À´Ï´Ù.
´ÙÄ«¶ó¹ÙÀÌ¿ÀÀÇ ½ÂÀξøÀÌ Á¦Ç°ÀÇ ÀçÆÇ • ¾çµµ, ÀçÆÇ • ¾çµµ¸¦ À§ÇÑ º¯Çü, »ó¿ë Á¦Ç°ÀÇ Á¦Á¶¿¡ »ç¿ëÇÏ´Â °ÍÀº ±ÝÁöµÇ¾î ÀÖ½À´Ï´Ù.
´ÙÄ«¶ó¹ÙÀÌ¿À Á¦Ç°¿¡ °ü·ÃµÈ ¶óÀ̼±½º¿Í ƯÇã¿¡ ´ëÇؼ­´Â ¶óÀ̼±½º ¸¶Å©¸¦ Ŭ¸¯ÇÏ¿© ³»¿ëÀ» È®ÀÎÇØ ÁֽʽÿÀ.
¶ÇÇÑ, ´Ù¸¥ Á¦Á¶¾÷ü Á¦Ç°¿¡ °üÇÑ ¶óÀ̼±½º ƯÇã¿¡ ´ëÇؼ­´Â °¢ Á¦Á¶¾÷üÀÇ À¥ »çÀÌÆ® ¶Ç´Â Á¦Á¶¾÷ü ¹ßÇàÀÇ Ä«Å¸·Î±×µî¿¡¼­ È®ÀÎÇØ ÁֽʽÿÀ.

Keyword :

Æ÷À¯·ù ¼¼Æ÷ ´Ü¹éÁú ¹ßÇö
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Suspension CHO cell¿ë ´ë·® ´Ü¹éÁú ¹ßÇö ½Ã½º..
CHOgro¢ç Expression System
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Suspension CHO cellÀ» »ç¿ëÇÑ ´Ü±â°£ Ç×ü/´Ü..
CHOgro¢ç High Yield Expression System
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´Ü¹éÁú disulfide BondÀÇ ÀçÁ¶ÇÕ¿¡
Corystein¢â£¨Purothionin£©
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¹«¼¼Æ÷ ´Ü¹éÁú ÇÕ¼º°è
Human Cell-Free Protein Expression System
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Human cell (HEK293)¿¡¼­ ´Ü¹éÁúÀ» °í¹ßÇö
pHEK293 Ultra Expression Vector