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huojinlong8610

½ð³æ (СÓÐÃûÆø)

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ΪÁË»ñµÃÖí°×ϸ°û½éËØ6£¨IL6£©ºÍ¦Á¸ÉÈÅËØ£¨IFN¦Á£©Ë«ÖØ»îÐÔµÄÈںϵ°°×£¬Ñо¿Æä×÷ΪÃâÒß×ô¼ÁµÄ¿ÉÐÐÐÔ£¬ÀûÓÃIL6ºÍIFN¦Á»ùÒòµÄ±àÂëÇøÐòÁÐÒÔ¼°Ô­ºË±í´ïÔØÌåpET32aÐòÁУ¬Éè¼ÆÁË´øÓÐÏÞÖÆÐÔøÇÐλµã¡¢LinkerÐòÁÐÒÔ¼°His±êÇ©µÄÌØÒìÐÔÒýÎïÀ©Ôö³öÁËÖíIL6ºÍIFN¦ÁµÄ³ÉÊìëÄ»ùÒòÐòÁУ¬²¢·Ö±ðÓë¿ËÂ¡ÔØÌåpMD18Á¬½Óºóת»¯E.coli DH5¦Á£¬ÌáÈ¡ÖÊÁ£Ã¸Çв¢Á¬½Ó¹¹½¨ÖØ×éÖÊÁ£pMD18-IL6-IFN¦Á£¬½«¸ÃÖØ×éÖÊÁ£ºÍ±í´ïÔØÌåpET32aͬʱøÇв¢Á¬½Ó¹¹½¨pET32-IL6-IFN¦ÁÖØ×éÖÊÁ££¬ÒÀ´Îת»¯E.coli DH5¦ÁºÍE.coli Rosetta (DE3)¸ÐÊÜ̬ϸ°û£¬²¢¾­²»Í¬Å¨¶ÈµÄIPTGÒÔ¼°²»Í¬Ê±¼ä½øÐÐÓÕµ¼±í´ï¡£½á¹û³É¹¦¹¹½¨ÁËIL6ºÍIFN¦ÁµÄÈںϱí´ïÔØÌ壬SDS-PAGE¼ì²âpET32-IL6-IFN¦ÁÈںϵ°°×ÔÚE.coli Rosetta (DE3)Öеõ½Á˽ϸ߱í´ï£¬Ä¿µÄµ°°×µÄÏà¶Ô·Ö×ÓÁ¿Îª44.96 KDa£¬ÓëÀíÂÛÔ¤ÆÚÖµÒ»Ö£¬³¬Éù²¨ºóSDS-PAGE¼ì²â·¢ÏÖµ°°×Ö÷ÒªÒÔ²»ÈÜÐԵİüº­ÌåÐÎʽ´æÔÚ¡£

To explore the feasibility of interleukin-6/interferon-¦Á (IL6/IFN¦Á) fusion protein as an immunoadjuvant, the coding sequences of IL6 and IFN¦Á were cloned. Considering the characteristics of these coding sequences and the sequence of prokaryotic expression vector pET32a, the mature peptide sequences of IL6 and IFN¦Á were amplified using the specific primers designed with restriction enzyme sites, linker sequences and His tag, and then transfected into E.coli DH5¦Á after linked with pMD18, respectively. The plasmids were extracted and digested by restriction enzyme respectively, and linked to construct the recombinant plasmid of pMD18-IL6-IFN¦Á. The plasmid was transformed into E.coli DH5¦Á and Rosetta (DE3), and then expressed under different concentrations' IPTG and times. The results showed that the recombinant plasmid of pET32-IL6-IFN¦Á was constructed successfully, SDS-PAGE results showed that recombinant fusion proteins were expressed highly in E.coli Rosetta (DE3), and the molecular weight of inclusion bodies is 44.96 KDa. pET32a-IL6-IFN¦Á was detected after ultrasonic treatment by PAGE and found existed as inclusion body form.

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ÒÑÔÄ   »Ø¸´´ËÂ¥   ¹Ø×¢TA ¸øTA·¢ÏûÏ¢ ËÍTAºì»¨ TAµÄ»ØÌû

yangruyiyantai

ľ³æ (ÖøÃûдÊÖ)

¡ï ¡ï ¡ï ¡ï ¡ï
°®ÓëÓêÏÂ: ½ð±Ò+1 2013-03-19 21:08:20
huojinlong8610: ½ð±Ò+5, ·­ÒëEPI+1, ûÓнøÐÐÈóÉ«£¬ÕÕ°áÁËÎÒÔ­À´µÄ 2013-03-20 15:33:23
phu_grassman: ½ð±Ò-1, ·­ÒëEPI-1, δ½øÐÐÈóÉ«£¬³·ÏúEPI¡£ 2013-04-04 09:20:15
To aquire the feasibility of interleukin-6/interferon-¦Á (IL6/IFN¦Á) fusion protein as an immunoadjuvant, the coding sequences of IL6 and IFN¦Á were cloned. Considering the characteristics of these coding sequences and the sequence of prokaryotic expression vector pET32a, the mature peptide sequences of IL6 and IFN¦Á were amplified using the specific primers designed with restriction enzyme sites, linker sequences and His tag, and then transfected into E.coli DH5¦Á after linked with pMD18, respectively.
The plasmids were extracted and digested by restriction enzyme respectively, and linked to construct the recombinant plasmid of pMD18-IL6-IFN¦Á. The plasmid was transformed into E.coli DH5¦Á and Rosetta (DE3), and then expressed under different concentrations' IPTG and times.
The results showed that the recombinant plasmid of pET32-IL6-IFN¦Á was constructed successfully, SDS-PAGE results showed that recombinant fusion proteins were expressed highly in E.coli Rosetta (DE3), and the relative molecular weight of inclusion bodies is 44.96 KDa.
pET32a-IL6-IFN¦Á was detected after ultrasonic treatment by PAGE and found existing as inclusion body form.
ÿÖÜÐĵû㱨¡£Ô´ÓÚÅ£½ò¡£ÖµµÃ½è¼ø¡£
2Â¥2013-03-19 17:27:45
ÒÑÔÄ   »Ø¸´´ËÂ¥   ¹Ø×¢TA ¸øTA·¢ÏûÏ¢ ËÍTAºì»¨ TAµÄ»ØÌû

cory0931

ÖÁ×ðľ³æ (ÎÄ̳¾«Ó¢)

Î×É½ÔÆ

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¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ¡ï ...
°®ÓëÓêÏÂ: ½ð±Ò+1 2013-03-19 21:08:24
huojinlong8610: ½ð±Ò+50, ¡ïÓаïÖú 2013-03-20 15:40:47
phu_grassman: ½ð±Ò+1, ·­ÒëEPI+1, thanks for your help/ 2013-04-04 09:21:21
СÊÔһϣ¬½ö¹©²Î¿¼£»
To obtain the fusion protein of swine interleukin-6 & interferon-¦Á(IL6 & IFN¦Á) and explore its feasibility of using as an immunoadjuvant, primers with restriction endonuclease sites, linker and his-tag was designed according to the sequences of Il6/IFN¦Á and vector pET32a. Matured peptide sequences of swine IL6 & IFN¦Áwere successfully cloned and amplified by transfecting into E. coli through pMD18, respectively. The pMD18 plasmids were extracted and enzymatically cut to build the recombinant plasmid pMD18-IL6-IFN¦Á(rpMD18-IL6-IFN¦Á), followed by transfection into competent E. coli DH5¦Á and E. coli Rosetta(DE3), and expressed by different IPTG concentration induction in different time. The results showed that the fusion expression plasmids were successfully built and highly expressed in E. coli Rosetta(DE3) according to SDS-PAGE detection. Molecular weight of the interest protein is 44.96kDa, which is agree with predicted value, and is mainly existed as insoluble inclusion bodies by SDS-PAGE detection after ultrasound treatment.
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3Â¥2013-03-19 19:34:18
ÒÑÔÄ   »Ø¸´´ËÂ¥   ¹Ø×¢TA ¸øTA·¢ÏûÏ¢ ËÍTAºì»¨ TAµÄ»ØÌû

hookhans

Ìú¸Ëľ³æ (ÖøÃûдÊÖ)

Farmer

ask  Rio2016.
where-there-is-a-will-there-is-a-way.
4Â¥2013-03-19 19:40:46
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Ïà¹Ø°æ¿éÌø×ª ÎÒÒª¶©ÔÄÂ¥Ö÷ huojinlong8610 µÄÖ÷Ìâ¸üÐÂ
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