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HL50285.5植物磷脂酶C试剂盒实验流程
点击次数:102 发布时间:2019-11-06

50285.5 v.A

 

ZHIWULINZHIMEIC(Phospholipase C)HUOXINGMEILIANXUXUNHUANFANYINGBISEFADINGLIANGJIANCESHIJIHE

CHANPINSHUOMINGSHU

 

ZHUYAOYONGTU

 

河北快3ZHIWULINZHIMEIC(Phospholipase C)HUOXINGMEILIANXUXUNHUANFANYINGBISEFADINGLIANGJIANCESHIJISHIYIZHONGZHIZAITONGGUOLINZHIMEIC、JIANXINGLINSUANMEI、DANJIANYANGHUAMEIHEGUOYANGHUAMEIMEILIANXUFANYINGXITONGZHONG,ZAILINZHIMEICYIZHIJIDECUNZAIXIA,SUOCHANSHENGDEHONGSECHANWUKUNYAANRANLIAO,ZAIFENGUANGGUANGDUYIXIAFENGZHIDEZENGGAO,JICAIYONGBISEFALAICEDINGZUZHILIEJIEYANGPINZHONGMEIHUOXINGDEERJINGDIANDEJISHUFANGFA。GAIJISHUJINGGUOJINGXINYANZHI、CHENGGONGSHIYANZHENGMINGDE。QISHIHEYUGEZHONGZHIWUZUZHI,BAOKUOZHONGZI(seed)、YEPIAN(leaf)、GEN(root)、PEITAIYE(cotyledon)、SHANGPEIZHOU(epicotyl)HUOWEILITIYANGPINDENGLINZHIMEICDEHUOXINGJIANCE。CHANPINYANGEWUJUN,JIDAOJIYONG,CAOZUOJIANJIE,XINGNENGWENDING。

 

JISHUBEIJING

 

LINZHIMEI(phospholipase),SHIYIZHONGZHIZHISHUIJIEMEI,CUIHUALINZHIDESHUIJIE,CHANSHENGZHIFANGSUANHEQITAQINZHICHANWU,CANYULINZHIDAIXIE。LINZHIMEIJIASHUYOUSIZHONGLEIXINGYAMEI,BAOKUOA、B、CHED。LINZHIMEIA(phospholipase A;PLA)ZHUYAOSHUIJIELINZHIsn-1(LINZHIMEIA1CUIHUA)HEsn-2(LINZHIMEIA2CUIHUA)WEIZHISHANGDEYIXIANJI;LINZHIMEIB(phospholipase B;PLB)TONGSHISHUIJIELINZHIDEsn-1HEsn-2WEIZHISHANGDEYIXIANJI,YOUCHENGWEIRONGXUELINZHIMEI(lysophospholipase);LINZHIMEIC(phospholipase C;PLC;EC3.1.4.3)ZAILINSUANJIQIAN(JILINSUANHEGANYOUZHIJIAN)SHUIJIE,SHIFANGERXIANJIGANYOU(diacylglycerol;DAG)HEDIERXINSHIFENZISANLINSUANJICHUN(inositol triphosphate);LINZHIMEID(phospholipase D;PLD)ZAILINSUANJIHOUSHUIJIE,SHIFANGLINZHISUAN(phosphatidic acid)。BURUDONGWULINZHIMEID,YOUCHENGWEILINZHIXIANDANJIANLINZHISUANSHUIJIEMEI(Phosphatidylcholine phosphatidohydrolase),YOU2GEYIGOUTI:LINZHIMEID1HED2,CUNZAIYUXIBAOZHIMOSHANG。LINZHIMEIC,YOUCHENGWEILUANLINZHIMEI(lecithinase)HUOα-DUSU(α-toxin),ZAIXIJUNPEIYANGYESHANGQINGZHONGCUNZAI,JUYOUSHUIJIESHENGWUMOSHANGDELUANLINZHI(lecithin),YINCIHUICHANSHENGRONGXUE、PIFUHUAISIXING(dermonecrotic)HEZHIMINGZUOYONG。LINZHIMEICHAIKEYISHUIJIEQIAOLINZHI(sphingomylin),NAOLINZHI(phosphatidylethanolamine)DENG。ZAIXIBAOXINHAOCHUANDAOTONGLUSHANGQIZHEZHONGYAOZUOYONG。ZHIWULINZHIMEICSHOUDAOGAILIZIDIAOJIE,ZAISHENGWUXINGHUOFEISHENGWUXINGYALIZHONGFAHUIZUOYONG。QICITONGGUODIAOJIEZHIZHIXINHAOTONGLUZHONG,KONGZHIHUAFENGUANSHENGZHANGHEZHIWUFAYU。JIYUDIWULUANLINZHI(lecithin),ZAILINZHIMEICXUANZEXINGYIZHIJIU73122CUNZAIYUFOUDEQINGKUANGXIA,TONGGUOLINZHIMEICDEZUOYONG,SHUIJIECHANSHENGWEIERXIANJIGANYOUHELINSUANDANJIAN(choline phosphate),JINERYOUJIANXINGLINSUANMEI(alkaline phosphatase)QULINSUANHUACHANSHENGDANJIAN(choline),HOUTONGGUODANJIANYANGHUAMEI(choline oxidase)HEGUOYANGHUAMEI(peroxidase)XITONG,CEDINGHONGSECHANWUKUNYAANRANLIAO(quinoneimine dye)DEFENGZHIBIANHUA(500nm BOZHANG),LAIDINGLIANGFENXILINZHIMEICDEHUOXING。QILIANXUXUNHUANFANYINGXITONGWEI:

 

PLC 

河北快3Leicithin  +  H2O     →      Diacylglycerol  +  Choline phosphate

 

 Alkaline phosphatase 

Choline phosphate  +  H2O     →      Choline  +  Phosphate

 

choline oxidase

Choline  +  O2  +  H2O  → Betaine aldehyde  +  2 H2O2

 

河北快3            peroxidase

河北快3 2 H2O2  +  4-Aminoantipyrine  +  Phenol     →    2 H2O  +   Quinoneimine dye   

 

CHANPINNEIRONG

 

河北快3QINGLIYE(Reagent A) 30HAOSHENG

河北快3LIEJIEYE(Reagent B) 10HAOSHENG

HUANCHONGYE(Reagent C)   5HAOSHENG

河北快3FANYINGYE(Reagent D) 600WEISHENG

河北快3MEICUYE(Reagent E) 600WEISHENG

DISEYE(Reagent F) 600WEISHENG

河北快3YINXINGYE(Reagent G) 600WEISHENG

河北快3ZHUANXINGYE(Reagent H) 100WEISHENG

河北快3CHANPINSHUOMINGSHU     1FEN

 

BAOCUNFANGSHI

 

BAOCUNQINGLIYE(Reagent A)ZAI4℃,QIYUDEBAOCUN-20℃BINGXIANGLI;DISEYE(Reagent F)BIMIANGUANGZHAO;YOUXIAOBAOZHENG6YUE

 

YONGHUZIBEI

 

1.5HAOSHENGLIXINGUAN:YONGYUYANGPINBAOCUNHEFANYINGCAOZUODERONGQI

15HAOSHENGZHUIXINGLIXINGUAN:YONGYUYANGPINZHIBEIDERONGQI

河北快3DOUNCEYUNJIANGQI:YONGYUZUZHIYUNHUA

河北快3(WEIXING)TAISHILIXINJI:YONGYUYANGPINCAOZUO

河北快3200WEISHENG1LIMIGUANGJINGBISEMINHUO96KONGBAN:YONGYUBISEDERONGQI

河北快3FENGUANGGUANGDUYIHUOMEIBIAOYI:YONGYUBISEFENXI

 

SHIYANBUZHOU

 

YI、YANGPINZHUNBEI

 

河北快31.ZHUNBEIHAOXINXIANDEZHIWUZUZHI(YEPIAN、GULI、ZHONGZIDENG),BINGCHENGZHONGYIQUEDING200ZHI500HAOKEZUZHIZHONGLIANG 

2.(XUANZEBUZHOU)FANGJINYULENGDE15HAOSHENGZHUIXINGLIXINGUAN

河北快33.(XUANZEBUZHOU)JIARU3HAOSHENGQINGLIYE(Reagent A)QINGXI1CI

河北快34.JIKEYONGDAOPIANQIESUIZUZHI

5.FANGJINYIGEYULENGDE15HAOSHENGZHUIXINGLIXINGUAN

6.JIARUYULENGDE1HAOSHENGLIEJIEYE(Reagent B)

河北快37.WOXUANZHENDANG5MIAO,CHONGFENHUNYUN

8.JIKEFANGJINYULENGDEDOUNCEYUNJIANGQI,ZAIBINGCAOLIYONGYUNJIANGBANGYUNHUAZUZHI(YUE80XIA)

9.JIANGSUOYOUZUZHIYUNJIANGWUYIRU1.5HAOSHENGLIXINGUAN,FANGJIN4℃WEIXINGTAISHILIXINJILIXIN10FENZHONG,SUDUWEI5000g(HUO7500RPM,LIRUeppendorf 5415)

10.XIAOXINYICHUSHANGQINGYE(ZHUYI:BUYAOCHUPENGCHENDIANWU)DAOLINGYIGEXINDEYULENGDE1.5HAOSHENGLIXINGUAN

河北快311.(XUANZEBUZHOU)RUGUOSHANGQINGYEHANYOUROUYANKEJIANDECANZHA,ZHONGFULIXIN5FENZHONGYICI,SUDUWEI5000g(HUO7500RPM,LIRUeppendorf 5415)

河北快312.FANGJIN4℃WEIXINGTAISHILIXINJILIXIN5FENZHONG,SUDUWEI16000g(HUO13000RPM,LIRUeppendorf 5415)

河北快313.XIAOXINYIQU1HAOSHENGSHANGQINGYEDAOXINDEYULENGDE1.5HAOSHENGLIXINGUAN

河北快314.(XUANZEBUZHOU)FANGJIN4℃CHAOSULIXINJILIXIN60FENZHONG,SUDUWEI105000g 

河北快315.(XUANZEBUZHOU)XIAOXINYIQU1HAOSHENGSHANGQINGYEDAOXINDEYULENGDE1.5HAOSHENGLIXINGUAN

河北快316.YIQU10WEISHENGJINXINGDANBAIDINGLIANGJIANCE(ZHUYI:JIANYISHIYONG BCADANBAIZHINONGDUDINGLIANGSHIJIHE-30031.1)

17.JIKEFANGJIN-70℃BAOCUNHUOZHIYUBINGCAOLIJIXUHOUXUCAOZUO

 

ER、CEDINGZHUNBEI

 

1.KAIQIBINGSHEDINGHAOFENGUANGGUANGDUYIHUOMEIBIAOYI(WENDUWEI37℃):BOZHANG500nm,BINGZHILING 

2.CONG-20℃BINGXIANGLIQUCHUSHIJI,ZHIYUBINGCAOLIRONGHUA;DISEYE(Reagent F)BIMIANGUANGZHAO

河北快33.HUANCHONGYE(Reagent C)SHIWENXIAJUNHENGWENDU

 

SAN、BEIJINGDUIZHAOCEDING

 

河北快31.YIQU170WEISHENGHUANCHONGYE(Reagent C)DAOXINDEBISEMINHUO96KONGBAN

2.JIARU25WEISHENGFANYINGYE(Reagent D)

3.JIARU10WEISHENGYINXINGYE(Reagent G)

河北快34.ZAI37℃WENDUXIAFUYU10FENZHONG

5.JIARU25WEISHENGMEICUYE(Reagent E)

河北快36.JIARU20WEISHENGDISEYE(Reagent F)

7.SHANGXIAQINGDAOSHUCI,HUNYUN 

8.ZAI37℃WENDUXIAFUYU30FENZHONG,BIMIANGUANGZHAO

河北快39.JIKEFANGJINFENGUANGGUANGDUYIHUOMEIBIAOYIJIANCE,CIWEIBEIJINGKONGDUIZHAO 

 

SI、YANGPINZONGHUOXINGCEDING

 

河北快31.YIQU170WEISHENGHUANCHONGYE(Reagent C)DAOXINDEBISEMINHUO96KONGBAN

河北快32.JIARU25WEISHENGFANYINGYE(Reagent D)

3.JIARU10WEISHENGDAICEYANGPIN(100WEIKEDANBAI)(ZHUYI:YANGPINXUQINGCHE)

4.WOXUANZHENDANG5MIAO

河北快35.ZAI37℃WENDUXIAFUYU10FENZHONG

6.JIARU25WEISHENGMEICUYE(Reagent E)

河北快37.JIARU20WEISHENGDISEYE(Reagent F)

8.SHANGXIAQINGDAOSHUCI,HUNYUN

河北快39.ZAI37℃WENDUXIAFUYU30FENZHONG,BIMIANGUANGZHAO

10.JIKEFANGJINFENGUANGGUANGDUYIHUOMEIBIAOYIJIANCE,CIWEIYANGPINZONGHUOXINGDUSHU  

 

河北快3WU、YANGPINFEITEYIHUOXINGCEDING

 

河北快31.YIQU160WEISHENGHUANCHONGYE(Reagent C)DAOXINDEBISEMINHUO96KONGBAN

河北快32.JIARU10WEISHENGZHUANXINGYE(Reagent H)

3.JIARU10WEISHENGDAICEYANGPIN(100WEIKEDANBAI)(ZHUYI:YANGPINXUQINGCHE)

河北快34.ZAI37℃WENDUXIAFUYU15FENZHONG

河北快35.JIARU25WEISHENGFANYINGYE(Reagent D)

6.ZAI37℃WENDUXIAFUYU10FENZHONG

7.JIARU25WEISHENGMEICUYE(Reagent E)

8.JIARU20WEISHENGDISEYE(Reagent F)

河北快39.SHANGXIAQINGDAOSHUCI,HUNYUN

河北快310.ZAI37℃WENDUXIAFUYU30FENZHONG,BIMIANGUANGZHAO

河北快311.JIKEFANGJINFENGUANGGUANGDUYIHUOMEIBIAOYIJIANCE,CIWEIYANGPINFEITEYIHUOXINGDUSHU

 

LIU、JISUANYANGPINHUOXING

 

(1)YANGPINZONGHUOXINGHEFEITEYIXINGHUOXING

 

BISEMINJISUAN

 

河北快3[(YANGPINDUSHU-BEIJINGDUSHU)X 0.25(TIXIRONGLIANG;HAOSHENG)XYANGPINXISHIBEISHU]÷[0.01(YANGPINRONGLIANG;HAOSHENG)X 12(HAOMOERXIGUANGXISHU)X 0.5 X 10(FANYINGSHIJIAN;FENZHONG)]=DANWEI/HAOSHENG 

 

河北快3ZHUANHUAN:DANWEI/HAOSHENG÷(YANGPINDANBAINONGDU)HAOKE/HAOSHENG=DANWEI/HAOKE

 

DANWEI=WEIMOERLIN/FENZHONG

 

96KONGBANJISUAN

 

[(YANGPINDUSHU-BEIJINGDUSHU)X 0.25(TIXIRONGLIANG;HAOSHENG)XYANGPINXISHIBEISHU]÷[0.01(YANGPINRONGLIANG;HAOSHENG)X 12(HAOMOERXIGUANGXISHU)X 0.5 X 0.6 (GUANGJINGJULI;LIMI)X 10(FANYINGSHIJIAN;FENZHONG)]=DANWEI/HAOSHENG 

 

ZHUANHUAN:DANWEI/HAOSHENG÷(YANGPINDANBAINONGDU)HAOKE/HAOSHENG=DANWEI/HAOKE

 

DANWEI=WEIMOERLIN/FENZHONG

 

(2)YANGPINTEYIHUOXING

 

YANGPINTEYIHUOXING=YANGPINZONGHUOXING-YANGPINFEITEYIHUOXING

 

ZHUYISHIXIANG

 

1.BENCHANPINWEI20CI(10GEYANGBEN)CAOZUO,BAOKUOBEIJINGDUIZHAO

2.CAOZUOSHI,XUDAISHOUTAO

河北快33.XITONGCAOZUOGUOCHENGZHONG,BEIJINGCEDINGZHIXU1CI

4.YANGPINXUCHENGQING,ZHIGUANZHONGYAO 

5.BISECEDINGHOU,BISEMINXUQINGXICHEDI

河北快36.YANGBENCEDINGDUSHUZENGGAO,BIAOMINGJUYOUMEIHUOXING

河北快37.JIANYIDAICEYANGBENDANBAINONGDUWEI100WEIKE/10WEISHENG;RUGUOYANGBENMEIHUOXINGGUODIHUOGUOGAO, ZEKEYIZENGJIAHUOJIANGDIYANGBENLIANG(BENGONGSITIGONG  BCADANBAIZHINONGDUDINGLIANGSHIJIHE-30031.1) 

8.RUGUODAICEYANGPINNONGDUGUOGAOHUOGUODI,KEYIDIAOZHENGYANGPINNONGDU

9.LINZHIMEICDANWEIHUOXINGDINGYIWEI:ZAI37℃,pH 7.3TIAOJIANXIA,MEIFENZHONGNEINENGGOUSHIFANG1WEIMOERLINSUOXUDEMEILIANGZUOWEIYIGEHUOXINGDANWEI

10.BENGONGSITIGONGXILIELINZHIMEIJIANCESHIJICHANPIN

 

ZHILIANGBIAOZHUN

 

1.BENCHANPINJINGJIANDINGXINGNENGWENDING

2.BENCHANPINJINGJIANDINGJIANCEMINGAN

 

SHIYONGCHENGNUO

 

BENGONGSIBINGZHE“XINYUZHISHANG、KEHUMANYI、ZHILIANGCHENGNUO”DEZONGZHIWEIWOMENDEYONGHUTIGONGCHANPINHEFUWU。YONGHUSHOUDAOHUOHOU,YINGANZHAOCHANPINSHUOMINGSHUSHANGDEGUIDINGTUOSHANBAOGUANBINGZAIYOUXIAOQINEISHIYONG。WOMENDECHANPINZAIXIAOSHOUQIANYIZUOYANGEDEZHILIANGJIANDING,BAOZHENGSHUOMINGSHUSUOSHUDESHIYONGXIAOGUO。ZAIHUODAOHOU10TIANNEIRUOFAXIANQUESHUBENCHANPINDEZHILIANGWENTI,QINGLIJIYISHUMIANXINGSHI(SHIYANBAOGAO,BINGJIANGGAICHANPINTUIHUI,JINGJIANYANQUEXICHANPINZHILIANGSUOZHI,BENGONGSIFUZEGENGHUANCHANPIN。RUXISHIYONGZHECUOWUCAOZUOSUOZHI,BENGONGSIBUCHENGDANYOUCIZAOCHENGDESUNSHI。

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