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cDNA Library Construction Kit
cDNA Library Construction Kit
Á¦Á¶»ç | Á¦Ç°ÄÚµå | Á¦Ç°¸í | ¿ë·® | °¡°Ý (ºÎ°¡¼¼º°µµ) |
ºñ°í |
---|---|---|---|---|---|
Clontech |
634901 |
SMART cDNA Library Construction Kit |
1 Set |
2,315,000¿ø | more |
Clontech |
635002 |
pRetro-Lib Vector |
20 ¥ìg |
684,000¿ø | more |
Clontech |
635003 |
pEXP-Lib Vector |
20 ¥ìg |
733,000¿ø | more |
- ÀûÀº ¾çÀÇ total RNA ¶Ç´Â poly A+ RNA·ÎºÎÅÍ °íÇ°ÁúÀÇ cDNA library Á¦ÀÛ
- ÃÖ¼Ò 50 ngÀÇ total RNA¿¡¼ cDNA ÇÕ¼º °¡´É
- µ¶Ã¢ÀûÀÎ SMART cDNA ÇÕ¼º¹ýÀ¸·Î full-length cDNA¸¦ Æ÷ÇÔÇÏ´Â library Á¦ÀÛ
- Adaptor ligation °úÁ¤ ºÒÇÊ¿ä
SMART cDNA Library Construction Kit´Â phage ¥ëTriplEx2 vector¿¡ full-length cDNA¸¦ cloningÇÏ´Â Á¦Ç°ÀÌ´Ù. ÀÌ Á¦Ç°Àº cDNAÁõÆøÀ» À§ÇÑ SMART ±â¼ú°ú adaptor ¾øÀÌ ¥ëTriplEx2 vector¿¡ cloningÇÏ´Â ±â¼úÀ» ÀÌ¿ëÇϸç ÃÊ±â ½Ã·á ¾ç¿¡ µû¶ó µÎ °¡Áö ½ÇÇè °úÁ¤À¸·Î ³ª´²Á® ÀÖ´Ù. ù ¹ø° °úÁ¤Àº »õ·Ó°Ô °í¾ÈµÈ long-distance PCR(LD-PCR) ¹æ¹ýÀ¸·Î 50 ngÁ¤µµÀÇ ÀûÀº ¾çÀÇ RNA ½Ã·á¿¡ Àû¿ëÇÒ ¼ö ÀÖ´Ù (1). µÎ ¹ø° °úÁ¤Àº Á» ´õ ÀϹÝÀûÀÎ °úÁ¤À¸·Î ºñ±³Àû ¸¹Àº ¾çÀÇ Ãʱ⠽÷á (¿¹¸¦ µé¸é, 1 ¥ìg ¶Ç´Â ±× ÀÌ»óÀÇ poly A+ RNA)À» »ç¿ëÇÏ´Â ½ÇÇèÀÚ¿¡°Ô ÀûÇÕÇÏ´Ù. µÎ °úÁ¤ ¸ðµÎ ¸¹Àº ¾çÀÇ full-lengthÀÇ double-strand cDNA¸¦ ÇÕ¼ºÇϱâ À§ÇÏ¿© 1st strand ÇÕ¼º°úÁ¤¿¡¼ SMART IV Oligonucleotide¸¦ ÀÌ¿ëÇÑ´Ù. µÎ ½ÇÇè °úÁ¤ ¸ðµÎ UserManual¿¡ Ç¥±âµÇ¾î ÀÖÀ¸¸ç °¢ÀÚÀÇ ½ÇÇè¿¡ ¸Â°Ô ¼±ÅÃÇϵµ·Ï ÇÑ´Ù. °¢°¢ÀÇ kit´Â cDNA library 7ȸ Á¦ÀÛ ºÐÀÌ´Ù.
¶ÇÇÑ, Æ÷À¯·ù ¹ßÇö¿ë vectorÀÎ pEXP-Lib, retroviral expression vectorÀÎ pRetro-Lib µî SMART cDNA Library Construction Kit¿Í ÇÔ²² »ç¿ëÇÒ ¼ö ÀÖ´Â 7°¡Áö vector°¡ º°µµ ÆǸŵǰí ÀÖ´Ù.
SMART library´Â ±âÁ¸ÀÇ ÀüÅëÀûÀÎ ¹æ¹ý¿¡ ºñÇØ ¸Å¿ì ³ôÀº È¿À²·Î Àüü±æÀ̸¦ Æ÷ÇÔÇÏ´Â cloneÀ» Á¦ÀÛÇÒ ¼ö ÀÖ´Ù. µû¶ó¼ SMART cDNA library¿¡¼ ¼±º°µÈ cloneÀº mRNAÀÇ 5¡¢-untranslated regionÀ» ¿ÏÀüÈ÷ Æ÷ÇÔÇϰԵȴ٠(2).
RNA/cDNA Quality Assay¸¦ »ç¿ëÇÏ¸é ½ÇÇèÀ» ½ÃÀÛÇϱâ Àü¿¡ Ãʱ⠽÷á·Î »ç¿ëµÇ´Â human RNAÀÇ »óŸ¦ È®ÀÎÇÒ ¼ö ÀÖ´Ù. º» ½ÇÇè¿¡´Â RT-PCR¹ýÀÌ ÀÌ¿ëµÇ¹Ç·Î ½ÇÇè¿¡ Àû¿ëÇÒ full-length cDNA°¡ ÁõÆøµÇ´ÂÁö ¿©ºÎ¸¦ Á÷Á¢ÀûÀ¸·Î È®ÀÎÇÒ ¼ö ÀÖ´Ù. ±âÁ¸ÀÇ ½ÇÇè½Ç¿¡¼ »ç¿ëÇÏ´ø ÀåºñµéÀ» ÀÌ¿ëÇϹǷΠºü¸£°Ô RNA »óŸ¦ È®ÀÎÇÒ ¼ö ÀÖÀ» »Ó¸¸ ¾Æ´Ï¶ó ºÒÆíÇÏ°í À¯ÇØÇÑ formaldehyde gelÀ» Á¦ÀÛÇÒ ÇÊ¿äµµ ¾ø´Ù.
Figure 1. Comparison of SMART cDNA synthesis vs. conventional synthesis for library construction.
Á¦Ç°¸í | Cat. No. | ¿ë·® | ¸Å´º¾ó | First Strand Synthesis | cDNA Amplification | Vector | Ư¡ |
SMART cDNA Library Construction Kit |
634901 | 10 ȸ | ¸Å´º¾ó | SMART IV Oligonucleotide | 5' PCR Primer | ¥ëTriplEx2 (lamda phage) |
* SMART cDNA ÇÕ¼º ±â¼ú * lamda phage¿¡ library Á¦ÀÛ * Á¦ÀÛµÈ library¸¦ °í°´ÀÌ ¿øÇÏ´Â plasmid·Î coversion±ÇÀåÇÏÁö ¾ÊÀ½ |
CDS III/3' PCR Primer | |||||||
In-Fusion SMARTer Directional cDNA Library Construction Kit |
634933 | 10 ȸ | ¸Å´º¾ó | SMARTer V Oligonucleotide | 5' PCR Primer II A | pSMART2IFD Linearized Vector |
* SMART±â¼ú In-Fusion Cloning ±â¼ú * ¿øÇÏ´Â vector¿¡ °ð¹Ù·Î library Á¦ÀÛ °¡´É(È¿À² °í·Á ÇÊ¿ä) * InsertÀÇ cloning ¹æÇ⼺ êó |
3¡¯ In-Fusion SMARTer CDS Primer | 3¡¯ In-Fusion SMARTer PCR Primer |
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