产品详情
pComb3XSS噬菌体展示质粒
基本信息
启动子: Lac/lac promoter
质粒大小: 4991bp
质粒标签: C-6×HIS，C-HA
原核抗性: 氨苄青霉素Amp
克隆菌株: 大肠杆菌Stbl3
培养条件: 37℃，有氧，LB
质粒简介
pComb3XSS是一个噬菌体展示质粒。pComb3X is the newest of the pComb vectors. Improvements over pComb3 include increased stability and introduction of an asymmetric SfiI cassette for directional cloning of full Fab, scFv, peptide and other protein for phage display. 6xHis and HA tags allow for purification and detection. An amber stop codon was introduced to turn-off expression of the pIII fusion protein by switching to a non-supressor strain of E. coli allowing production of soluble protein without subcloning. Alternatively, the gene for phage protein pIII can be removed by SpeI/NheI digest. pComb3XSS is recommended for preparation of vector for library cloning. The “SS” refers to the double stuffer, a 1200bp stuffer in the Fab light chain cloning region bounded by SacI and XbaI restriction sites and a 300bp stuffer in Fab heavy chain cloning region bound by XhoI and SpeI restriction sites. Also, the 1600bp double stuffer (both stuffers plus the leader sequence between the Fab light chain and heavy chain cloning regions) can be removed by SfiI digest so that non-Fab genes of interest can be cloned. Also available on Addgene: pComb3XTT and pComb3XLambda are only needed at templates for the construction of chimeric Fab libraries as described in Phage Display: A Laboratory Manual. pComb3XTT can also be used as an Fab expression control. 3rd generation plasmid for phage display on modified geneIII, contains stuffer fragment.
质粒图谱
pComb3XSS噬菌体展示质粒使用说明：
1、收到质粒干粉后请先5000rpm离心1min，再加入20μl无菌水溶解质粒，室温放置1min；
2、从-80℃冰箱中取出相应的感受态，置于冰盒上解冻，并做好标记；
3、取2μl质粒加至100μl感受态中，冰浴30min；
4、42℃热激90s，再冰浴2min；
5、加入900μl无抗的LB液体培养基，180rpm震荡培养45min；
6、6000rpm离心5min，仅留100ul上清混匀菌体沉淀；
7、混匀后的菌液加至对应抗性的LB平板上，倒入适量玻璃珠，涂匀液体；
8、将平板正向培养1h，再倒置培养12h~16h；
9、挑取单克隆菌落至对应抗性的LB液体培养基中，震荡培养12h~16h，根据实验需要提取质粒。
pComb3XSS噬菌体展示质粒注意事项：
1、如果您收到的是甘油菌种，请先四区划线，挑取单克隆培养。
2、如果第二天转化平板长的过多，请将质粒按比例稀释后再转化。
