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TaKaRa
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  • Å䡤ÀÏ¿äÀÏ, °øÈÞÀÏÀº ÈÞ¹«ÀÔ´Ï´Ù.
Cloned

SP6 RNA Polymerase

-

Á¦Á¶»ç Á¦Ç°ÄÚµå Á¦Ç°¸í ¿ë·® °¡°Ý
(ºÎ°¡¼¼º°µµ)
ºñ°í »ç¿ëÀڸŴº¾ó
Takara
2520A
SP6 RNA Polymerase
°ü·ÃÇмú ±¸¸ÅÇϱâ
3,000 U
185,600¿ø 
232,000¿ø
°¡°ÝÇÒÀÎ
09.02 ~ 10.31
2520A_DS.v2107Da.pdf
Takara
2520B
SP6 RNA Polymerase
°ü·ÃÇмú ±¸¸ÅÇϱâ
15,000 U (2520A x 5)
1,050,000¿ø 

Á¦Ç°¼³¸í
º» È¿¼Ò´Â SP6 promoter¸¦ Æ÷ÇÔÇÑ ÀÌÁß °¡´Ú DNA¸¦ ÁÖÇü, NTP¸¦ ±âÁú·Î »ç¿ëÇÏ¿© promoter ÇÏ·ùÀÇ ÁÖÇü DNA¿¡ »óº¸ÀûÀÎ ´ÜÀÏ °¡´Ú RNA¸¦ ÇÕ¼ºÇÑ´Ù. SP6 promoter ¼­¿­¿¡ ³ôÀº ƯÀ̼ºÀ» º¸ÀÌ°í ´Ù¸¥ »ý¹° À¯·¡ÀÇ promoter´Â ÀνÄÇÏÁö ¾Ê´Â´Ù.
º¸Á¸
-20¡É
³óµµ
50 U/§¡
Çü»ó

10 mM

Potassium Phosphate (pH7.9)

1 mM

DTT

0.1 mM

EDTA

150 mM

NaCl

50%

Glycerol

÷ºÎ buffer Á¶¼º (10x)

400 mM

Tris-HCl (pH7.5)

60 mM

MgCl2

20 mM

Spermidine

100 mM

DTT [º°Ã·]

0.1%

BSA [º°Ã·]

È°¼ºÀÇ Á¤ÀÇ
37¡É, pH7.5¿¡¼­ 1 ½Ã°£ µ¿¾È 1 nmolÀÇ [3H]GMP¸¦ »ê¼ºÀÇ Ä§Àü¹°·Î(acid-insoluble products)·Î ÇÕ¼ºÇÏ´Â È¿¼ÒÈ°¼ºÀ» 1 UÀ¸·Î ÇÑ´Ù.
È°¼º ÃøÁ¤¿ë ¹ÝÀÀ¾× Á¶¼º

40 mM

Tris-HCl (pH7.5)

6 mM

MgCl2

10 mM

DTT

2 mM

Spermidine

°¢ 0.5 mM

ATP, UTP, CTP

0.5 mM

[3H]GTP

0.01%

BSA

1 §¶/100 §¡

pSP65 DNA

»ç¿ë»óÀÇ ÁÖÀÇ
- ÃÖÀû pH´Â 7.5ÀÌ´Ù.
- Mg2+¿Í ȯ¿øÁ¦¸¦ ÇÊ¿ä·Î ÇÏ°í, BSA ¹× SpermidineÀÇ Ã·°¡¿¡ ÀÇÇØ È°¼ºÈ­µÈ´Ù.
- »ç¿ë½Ã¿¡´Â ÷ºÎ buffer¿¡ ¹Ýµå½Ã DTT¿Í BSA¸¦ ÷°¡ÇÑ´Ù. ¹ÌÆ÷ÇÔ ½Ã È°¼ºÀÌ ³ªÅ¸³ªÁö ¾ÊÀ» ¼ö ÀÖ´Ù.
- ÃÖÀû ¿Âµµ´Â 40¡É (37¡ÉÀÇ ¾à 130% È°¼º)¿¡¼­ È¿¼Ò·®Àº 300 U/§¢±îÁö, ÁÖÇü·®Àº 100 §¶/§¢±îÁö ÇÕ¼º·®ÀÌ Á¤ºñ·ÊÇÏ°Ô µÈ´Ù. ¹ÝÀÀ½Ã°£ÀÌ 1½Ã°£À» ³Ñ´Â °æ¿ì 37¡É¿¡¼­ ¹ÝÀÀÀ» ÃßõÇÏ´Ù.
- ƯÁ¤ ¿µ¿ª¸¸ÀÇ Àü»ç È¿À²À» ³ôÀÌ·Á¸é, ±× ¿µ¿ªÀÇ ÇÏ·ùÀÇ ÁÖÇü DNA¸¦ ÆòÈ°¸»´Ü (blunt end) ¶Ç´Â 5¡¯-µ¹Ã⸻´Ü(5¡¯-overhang)À¸·Î Àý´ÜÇÏ´Â °ÍÀ» ÃßõÇÑ´Ù.
¿ëµµ
- °­ÇÏ°Ô Ç¥½ÄµÈ RNA probe Á¦ÀÛ
- RNA splicing ¹ÝÀÀ Àü±¸Ã¼ Á¦ÀÛ
- Cap analog¸¦ primer·Î ÀÌ¿ëÇÑ capped mRNAÀÇ Á¦ÀÛ
±â¿ø
Escherichia coli carrying the plasmid containing phage SP6 RNA polymerase gene
ÀϹÝÀû ¼ºÁú
- ºÐÀÚ·®: ¾à 96,000
- Subunit: single stranded polypeptide
- ÃÖÀû pH: pH7.5
- º¸Á¶ÀÎÀÚ
   Mg2+ (ÃÖÀû³óµµ: 6 mM)
   ȯ¿øÁ¦ (10 mM DTT ¶Ç´Â 20 mM 2-mercaptoethanol)
- È°¼ºÈ­Á¦: 2 mM spermidine (4 mM¿¡¼­´Â DNA¿Í °øħ)
- ÀúÇØÁ¦: MEM (10-4 M), pCMB (5X10-8 M)¿¡¼­ ºÒÈ°¼ºÈ­
»ó´ë È°¼º
- ¹ÝÀÀ ¿Âµµ¿Í »ó´ë È°¼º (37¡ÉÀ» 100%·Î ÇßÀ» ¶§)
¹ÝÀÀ ¿Âµµ (¡É)

20

25

37

40

45

»ó´ë È°¼º (%)

0

10

100

130

80


- pH¿Í »ó´ëÈ°¼º
pH

7.5

7.7

7.8

8.0

»ó´ë È°¼º (%)

100

75

33

4


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0.5-10 kb ssRNA Ladder Marker