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RNAÀÇ 5¡¯ ¸»´ÜÀ» Cap 0 ¡æ Cap 1·Î º¯È¯ÇÏ´Â È¿¼Ò

mRNA Cap 2-O-Methyltransferase

-

Á¦Á¶»ç Á¦Ç°ÄÚµå Á¦Ç°¸í ¿ë·® °¡°Ý
(ºÎ°¡¼¼º°µµ)
ºñ°í »ç¿ëÀڸŴº¾ó
Takara
2470A
mRNA Cap 2-O-Methyltransferase
°ü·ÃÇмú ±¸¸ÅÇϱâ
2,500 U
114,000¿ø  2470A_DS.v2208Da.pdf
Takara
2470B
mRNA Cap 2-O-Methyltransferase
°ü·ÃÇмú ±¸¸ÅÇϱâ
10,000 U (2470A x 4)
412,000¿ø 

  • 7-methylguanylate(m7G) cap ±¸Á¶ (Cap 0)¸¦ °¡Áø RNAÀÇ Ã¹¹ø° nucleotide 2’-O ºÎÀ§¸¦ ƯÀÌÀûÀ¸·Î ¸ÞÆ¿È­
  • mRNAÀÇ ¸é¿ª¿ø¼º °¨¼Ò
  • mRNAÀÇ translation È¿À² Áõ°¡
Á¦Ç°¼³¸í
mRNA Cap 2'-O-Methyltransferase (2’-O-MTase)´Â vaccinia ¹ÙÀÌ·¯½º À¯·¡ÀÇ methyltransferase·Î 5’ ¸»´Ü¿¡ Cap 0 ±¸Á¶ (7-methylguanylate(m7G))¸¦ °¡Áø RNA¿¡ ù¹ø° nucleotideÀÇ 2’-O ºÎÀ§¸¦ ƯÀÌÀûÀ¸·Î ¸ÞÆ¿È­ÇÏ¿© Cap 1ÀÇ ±¸Á¶·Î ¸¸µç´Ù. RNA¿¡¼­ Cap 1ÀÇ ±¸Á¶´Â ü³»¿¡¼­ ¸é¿ª¿ø¼ºÀ» °¨¼Ò½ÃÅ°°í translation È¿À²À» Áõ°¡½ÃÅ°´Â °ÍÀ¸·Î ¾Ë·ÁÁ® ÀÖ´Ù.
ÀÌ Á¦Ç°¿¡´Â ¹ÝÀÀ buffer¿Í ¸ÞÆ¿±âÀÇ °ø¿©Ã¼ S-adenosylmethionine (SAM)ÀÌ Æ÷ÇԵǾî ÀÖ¾î °íÈ¿À²·Î Cap 0 RNA¿¡¼­ Cap 1 RNA¸¦ Á¦ÀÛÇÒ ¼ö ÀÖ´Ù.
¶ÇÇÑ ÀÌ È¿¼Ò´Â Vaccinia Capping Enzyme (Code 2460A)¿Í °°ÀÌ »ç¿ëÇÏ¿© uncapped RNA (5’-triphosphate RNA)·ÎºÎÅÍ Cap 1ÀÇ ±¸Á¶¸¦ °¡Áø RNA¸¦ one-stepÀ¸·Î Á¦ÀÛÇÒ ¼ö ÀÖ´Ù (»ç¿ëÀÚ ¸Þ´º¾ó protocol-2 Âü°í).
¿ëµµ
- Vaccinia Capping Enzyme (VCE) µîÀ¸·Î m7G cap±¸Á¶ (Cap 0)°¡ ºÎ°¡µÈ RNA¸¦ Cap 1 ±¸Á¶·Î º¯È¯.
- One-stepÀ¸·Î uncapped RNA (5’-triphosphate RNA)¿¡ Cap 1ÀÇ ±¸Á¶ ºÎ¿© (¡Øº°µµÀÇ VCE ÇÊ¿ä).
- in vitro transcription (IVT) µîÀ¸·Î ÇÕ¼ºÇÑ m7G cap±¸Á¶ (Cap 0)¸¦ °®´Â RNA¸¦ Cap 1 ±¸Á¶·Î º¯È¯ (ARCA (anti reverse cap analog) »ç¿ë ½Ã).
- mRNAÀÇ in vivo Àû¿ë ½Ã translation È¿À² Áõ°¡ ¹× ¸é¿ª¿ø¼º °¨¼Ò
³»¿ë

2,500 U

 

mRNA Cap 2’-O-Methyltransferase (50 U/§¡)

100 §¡

 

10× Capping Buffer

100 §¡

 

S-adenosylmethionine (SAM) (32 mM)

º¸Á¸ -20¡É
³óµµ 50 U/§¡
±â¿ø
Escherichia coli carrying a plasmid containing the gene for vaccinia virus mRNA cap 2’-O-methyltransferase
ÀϹÝÀûÀÎ ¼ºÁú
- Áú·®: ¾à 40 kDa
È°¼ºÀÇ Á¤ÀÇ
37¡É¿¡¼­ 1½Ã°£µ¿¾È 10 pmolÀÇ Cap 0 RNA¸¦ ¸ÞÆ¿È­ÇÏ´Â È¿¼Ò ¾çÀ» 1UÀ¸·Î ÇÑ´Ù.
È°¼º ÃøÁ¤¿ë ¹ÝÀÀÁ¦ Á¶¼º

 

Capping Buffer

1 mM

 

DTT

13.75 μCi/§¢

 

[3H]SAM

2 §¶/50 §¡

 

100 nt Cap 0 RNA

»ç¿ë»óÀÇ ÁÖÀÇ
- È¿¼ÒÀÇ °­ÇÑ ±³¹ÝÀº »ï°¡ÇÑ´Ù.
- Template RNA ¹× ½ÇÇè ±â±¸¿¡ RNase°¡ ¿À¿°µÈ °æ¿ì RNAÀÇ yield°¡ ³·¾ÆÁö°Å³ª fragmentation µÈ´Ù. ¹ÝÀÀ¿¡ »ç¿ëÇÏ´Â Æ©ºê, micropipette tip µîÀ» RNA½ÇÇè Àü¿ëÀ¸·Î »ç¿ëÇÏ°í ÀÏȸ¿ë Àå°©À» »ç¿ëÇÏ¿© RNaseÀÇ ¿À¿°ÀÌ ÀϾÁö ¾Êµµ·Ï ÁÖÀÇÇÑ´Ù.
½ÇÇè ¿¹½Ã
- »ç¿ëÀÚ ¸Þ´º¾ó Âü°í
°ü·ÃÁ¦Ç°
- uncapped RNAÀÇ 5’ ¸»´Ü¿¡ Cap 0 ±¸Á¶¸¦ ºÎ¿©ÇÏ´Â È¿¼Ò Vaccinia Capping Enzyme (Code 2460A)
- °íÇ°Áú °í¼öÀ² in vitro transcription ½Ã½ºÅÛ Takara IVTpro™ mRNA Synthesis System (Code 6141)
- Transfection ½Ã¾à Àü¹® ºê·£µå MirusÀÇ mRNA Àü¿ë Transfection ½Ã¾à TransIT-mRNA Transfection Kit (Code MIR 2250)
- ½ÇÇè ±â±â¿¡¼­ RNase ¿À¿° Á¦°Å RNase-OFF® (RNase Contamination Á¦°Å ¿ë¾×) (Code 9037)

Keyword :

in vitro transcription
>
°íÇ°Áú °í¼öÀ² in vitro transcriptionÀ» À§ÇÑ ..
Takara IVTpro¢â mRNA Synthesis System
>
dsRNA »ý¼ºÀÌ Àû¾î ÀÓ»óºÐ¾ßÀÇ ¿¬±¸°³¹ß¿¡ ÀûÇÕ..
PrimeCap¢â T7 RNA Polymerase (low dsRNA)
>
Poly(A) ÀÌÈÄ Ãß°¡¼­¿­ÀÌ ¾ø´Â scarless mRNA ..
Template Vector (BspQ I) for T7 mRNA Synthesis
>
RNAÀÇ 5¡¯-¸»´Ü¿¡ Cap (Cap 0)À» ºÎ°¡ÇÏ´Â È¿¼Ò
Vaccinia Capping Enzyme
>
RNAÀÇ 5¡¯ ¸»´ÜÀ» Cap 0 ¡æ Cap 1·Î º¯È¯ÇÏ´Â ..
mRNA Cap 2-O-Methyltransferase
>
´Ù¾çÇÑ ¿Âµµ¿¡¼­ ¹ÝÀÀÇÒ ¼ö ÀÖ´Â Capping enzym..
Faustovirus Capping Enzyme (S17)
>
Poly(A) ¹× ¹Ýº¹ ¼­¿­À» Æ÷ÇÔÇÏ´Â À¯ÀüÀÚÀÇ Å¬..
Takara Stable Competent Cells
>
RNA ÇÕ¼º ¼öÀ²ÀÌ Çâ»óµÈ
T7 RNA Polymerase ver.2.0
>
Àú³óµµ DTT¿¡¼­µµ ¾ÈÁ¤ÀûÀÎ RNase inhibitor
Recombinant RNase Inhibitor ver.2.0
>
DNA, RNA ÇÕ¼º ¹ÝÀÀÀ» ÃËÁø
Pyrophosphatase (inorganic)
>
IVT Àû¿ë¿¡ ÃÖÀû, Type IIS Á¦ÇÑÈ¿¼Ò
BspQ I
>
´Ù¾çÇÑ Å©±âÀÇ ssRNA »çÀÌÁî È®ÀÎ
0.5-10 kb ssRNA Ladder Marker
>
BspQ IÀ» »ç¿ëÇÑ °í¼öÀ² IVT ½Ã½ºÅÛ
(BspQ I) IVTpro¢â ½Ã½ºÅÛ
>
Low dsRNA Polymerase¸¦ »ç¿ëÇÑ °í¼öÀ² IVT ½Ã..
(PrimeCap¢â T7) IVTpro¢â ½Ã½ºÅÛ