基本信息
名称：pComb3XLambda噬菌体展示质粒
别称: Plasmid#63892

启动子:	Lac/lac promoter
复制子:	pUC
质粒大小:	4772bp
质粒标签:	C-6×His，C-HA
原核抗性:	Amp
克隆菌株:	TG1
培养条件:	37度

质粒属性

载体宿主：	大肠杆菌,噬菌体
载体用途：	蛋白表达,抗体表达
基因种属：	空载体
基因类型：	ORF
原核抗性：	Amp
真核抗性：
荧光蛋白：

质粒简介
Backbone pComb3X containing PCR template for human antibody lambda light chain constant region.
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. pComb3XLambda can be used as a template to amplify the lambda light chain constant region for construction of chimeric antibody libraries as described in:
Barbas, C. F., III; Burton, D. R.; Scott, J.K., Silverman, G.J. Eds. (2001) Phage Display: A Laboratory Manual; Cold Spring Harbor Laboratory Press: Cold Spring Harbor, New York
Please note that there is a A246V mutation in heavy chain region; however, this vector is meant to be used as a PCR template for the light chain region from base# 613-1016.

本质粒参考文献：Methods for the generation of chicken monoclonal antibody fragments by phage display.

质粒图谱


质粒序列
LOCUS Exported 4772 bp ds-DNA circular SYN 13-OCT-2017
DEFINITION synthetic circular DNA
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 4772)
AUTHORS .
TITLE Direct Submission
http://www.miaolingbio.com
FEATURES Location/Qualifiers
source 1..4772
/organism="synthetic DNA construct"
/mol_type="other DNA"
protein_bind 109..130
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence 
of cAMP."
promoter 145..175
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 183..199
/label=lac operator
/bound_moiety="lac repressor encoded by lacI"
/note="The lac repressor binds to the lac operator to 
inhibit transcription in E. coli. This inhibition can be 
relieved by adding lactose or 
isopropyl-beta-D-thiogalactopyranoside (IPTG)."
CDS 1743..1760
/codon_start=1
/product="6xHis affinity tag"
/label=6xHis
/translation="HHHHHH"
CDS 1767..1793
/codon_start=1
/product="HA (human influenza hemagglutinin) epitope tag"
/label=HA
/translation="YPYDVPDYA"
rep_origin complement(2387..2842)
/direction=LEFT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow 
indicates direction of (+) strand synthesis"
promoter 2869..2973
/gene="bla"
/label=AmpR promoter
CDS 2974..3834
/codon_start=1
/gene="bla"
/product="beta-lactamase"
/label=AmpR
/note="confers resistance to ampicillin, carbenicillin, and
related antibiotics"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
LIKHW"
rep_origin 4005..4593
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
replication"
ORIGIN
1 agagcgccca atacgcaaac cgcctctccc cgcgcgttgg ccgattcatt aatgcagctg
61 gcacgacagg tttcccgact ggaaagcggg cagtgagcgc aacgcaatta atgtgagtta
121 gctcactcat taggcacccc aggctttaca ctttatgctt ccggctcgta tgttgtgtgg
181 aattgtgagc ggataacaat tgaattcagg aggaatttaa aatgaaaaag acagctatcg
241 cgattgcagt ggcactggct ggtttcgcta ccgtggccca ggcggccgag ctcgtgctga
301 ctaacgccct cagtgtctgg ggccccaggg cagagggtca ccatctcctg cactgggagc
361 agctccaacc tcggggcagc ttatgatgta cactggtacc accaggttcc aggaacggcc
421 cccaaactcc tcctctatga taacatcaag cggccctcag gggtccctga ccgattctct
481 ggctccaagt ctggcacctc agcctccctg gccatcactg ggctccaggc tgaggacgag
541 gctgattatt actgcaacag agtaacagct ggcctcacac gttcggcgga gggaccaagc
601 tgaccgtcct aagtcagccc aaggctgccc cctcggtcac tctgttcccg ccctcctctg
661 aggagcttca agccaacaag gccacactgg tgtgtctcat aagtgacttc tacccgggag
721 ccgtgacagt ggcctggaag gcagatggca gccccgtcaa ggcgggagtg gagaccacca
781 caccctccaa acaaagcaac aacaagtacg cggccagcag ctatctgagc ctgacgcctg
841 agcagtggaa gtcccacaga agctacagct gccaggtcac gcatgaaggg agcaccgtgg
901 agaagacagt ggcccctaca gaatgttcat aattctagat aattaattag gaggaattta
961 aaatgaaata cctattgcct acggcagccg ctggattgtt attactcgct gcccaaccag
1021 ccatggccga ggtgcagctg ctcgagcagt ctggggctga ggtgaagaag cctgggtcct
1081 cggtgaaggt ctcctgcagg gcttctggag gcaccttcaa caattatgcc atcagctggg
1141 tgcgacaggc ccctggacaa gggcttgagt ggatgggagg gatcttccct ttccgtaata
1201 cagcaaagta cgcacaacac ttccagggca gagtcaccat taccgcggac gaatccacgg
1261 gcacagccta catggagctg agcagcctga gatctgagga cacggccata tattattgtg
1321 cgagagggga tacgattttt ggagtgacca tgggatacta cgctatggac gtctggggcc
1381 aagggaccac ggtcaccgtc tccgcagcct ccaccaaggg cccatcggtc ttccccctgg
1441 caccctcctc caagagcacc tctgggggca cagcggccct gggctgcctg gtcaaggact
1501 acttccccga accggtgacg gtgtcgtgga actcaggcgc cctgaccagc ggcgtgcaca
1561 ccttcccggc tgtcctacag tcctcaggac tctactccct cagcagcgtg gtgaccgtgc
1621 cctccagcag cttgggcacc cagacctaca tctgcaacgt gaatcacaag cccagcaaca
1681 ccaaggtgga caagaaagtt gagcccaaat cttgtgacaa aactagtggc caggccggcc
1741 agcaccatca ccatcaccat ggcgcatacc cgtacgacgt tccggactac gcttcttagg
1801 agggtggtgg ctctgagggt ggcggttctg agggtggcgg ctctgaggga ggcggttccg
1861 gtggtggctc tggttccggt gattttgatt atgaaaagat ggcaaacgct aataaggggg
1921 ctatgaccga aaatgccgat gaaaacgtgc tacagtctga cgctaaaggc aaacttgatt
1981 ctgtcgctac tgattacggt gctgctatcg atggtttcat tggtgacgtt tccggccttg
2041 ctaatggtaa tggtgctact ggtgattttg ctggctctaa ttcccaaatg gctcaagtcg
2101 gtgacggtga taattcacct ttaatgaata atttccgtca atatttacct tccctccctc
2161 aatcggttga atgtcgccct tttgtctttg gcgctggtaa accatatgaa ttttctattg
2221 attgtgacaa aataaactta ttccgtggtg tctttgcgtt tcttttatat gttgccacct
2281 ttatgtatgt attttctacg tttgctaaca tactgcgtaa taaggagtct taagctagct
2341 aattaattta agcggccgca gatctgctct ctgaggagga tctgggaaat tgtaagcgtt
2401 aatattttgt taaaattcgc gttaaatttt tgttaaatca gctcattttt taaccaatag
2461 gccgaaatcg gcaaaatccc ttataaatca aaagaataga ccgagatagg gttgagtgtt
2521 gttccagttt ggaacaagag tccactatta aagaacgtgg actccaacgt caaagggcga
2581 aaaaccgtct atcagggcga tggcccacta cgtgaaccat caccctaatc aagttttttg
2641 gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag ggagcccccg atttagagct
2701 tgacggggaa agccggcgaa cgtggcgaga aaggaaggga agaaagcgaa aggagcgggc
2761 gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa ccaccacacc cgccgcgctt
2821 aatgcgccgc tacagggcgc gtcaggtggc acttttcggg gaaatgtgcg cggaacccct
2881 atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga
2941 taaatgcttc aataatattg aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc
3001 cttattccct tttttgcggc attttgcctt cctgtttttg ctcacccaga aacgctggtg
3061 aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg gttacatcga actggatctc
3121 aacagcggta agatccttga gagttttcgc cccgaagaac gttttccaat gatgagcact
3181 tttaaagttc tgctatgtgg cgcggtatta tcccgtattg acgccgggca agagcaactc
3241 ggtcgccgca tacactattc tcagaatgac ttggttgagt actcaccagt cacagaaaag
3301 catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac catgagtgat
3361 aacactgcgg ccaacttact tctgacaacg atcggaggac cgaaggagct aaccgctttt
3421 ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa
3481 gccataccaa acgacgagcg tgacaccacg atgcctgtag caatggcaac aacgttgcgc
3541 aaactattaa ctggcgaact acttactcta gcttcccggc aacaattaat agactggatg
3601 gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg ctggtttatt
3661 gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc actggggcca
3721 gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc aactatggat
3781 gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg gtaactgtca
3841 gaccaagttt actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg
3901 atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg
3961 ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt
4021 ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg
4081 ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata
4141 ccaaatactg tccttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca
4201 ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag
4261 tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc
4321 tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga
4381 tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg
4441 tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac
4501 gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg
4561 tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg
4621 ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc ccctgattct
4681 gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag ccgaacgacc
4741 gagcgcagcg agtcagtgag cgaggaagcg ga
//

质粒菌株产品操作说明书
一、扩增流程
收到产品后，请先根据产品管壁标签来判断产品形式，并在扩增前准确查找该质粒菌株的抗性、感受态和培养温度。
1、质粒干粉（常温运输，存于-20度，90天保质期，请务必转化挑单克隆培养，不要直接使用和测序）
①收到质粒干粉后请先5000rpm离心1min，再加入20μl ddH2O去离子水溶解质粒；
②取1支100μl 感受态于冰上解冻10min，加入2μl质粒，再冰浴30min后，42℃热激60s，不要搅动，再冰浴2min；（从第二步开始均要在超净工作台中无菌操作）
③加入900μl无抗的LB液体培养基，180rpm震荡37℃培养45min；
④6000rpm离心5min，仅留100μl上清液重悬细菌沉淀，并涂布至目标质粒抗性的LB平板上；（可使用本平台的平板涂布专用玻璃珠进行涂布，可以比传统涂布方法获得更多转化子）
⑤将平板正向培养1h，再倒置37℃培养14h。如果要求是30度则培养20h；
（菌落过多则将质粒稀释后再转化。没有菌落则加入10μl质粒转化。另不要直接转表达感受态，要先转克隆感受态，重提质粒后再导入表达感受态）
⑥挑取单菌落至LB液体培养基中，加入对应抗生素，220rpm震荡培养14h，根据实验需要和质粒提取试剂盒说明书提取质粒。
2、甘油菌种（冰袋运输，存于-80℃，保质期90天，请务必划线挑单克隆培养）
四区划线后挑单菌落培养，酵母菌需要先液体复苏再四区划线，再挑单菌落液体培养。
3、穿刺菌种（冰袋运输，存于4℃，保质期7天）
穿刺接种，液体培养后四区划线，再挑单菌落液体培养。
4、菌落平板（冰袋运输，存于4℃，保质期7天）
直接挑取单菌落至液体培养基中。
5、液体质粒（冰袋运输，存于-20℃，保质期90天）
单独提取的液体质粒收到后可直接使用。
6、滤纸质粒（常温运输，存于-20度，90天保质期，请务必转化挑单克隆培养，不要直接使用和测序）
收到货后将滤纸画圈部分剪下放入EP管中，加100ul无菌水将滤纸浸湿并浸泡5min，吸取5ul质粒转化，离心全涂。

二、转化图片

P1880/pENTR223-ICMT人源基因质粒

P1895/pENTR223-PIP4K2B人源基因质粒

P1896/pENTR223-LOC142937人源基因质粒

P1897/pENTR223-INIP人源基因质粒

P1898/pENTR223-SPCS1人源基因质粒

P1899/pENTR223-ANAPC16人源基因质粒

P1900/pENTR223-BLOC1S2人源基因质粒

P1901/pENTR223-MGST1人源基因质粒

P1925/pENTR223-GLIPR1L1人源基因质粒

P1926/pENTR223-GINS4人源基因质粒

P1927/pENTR223-DUSP19人源基因质粒

P1928/pENTR223-PRDX1人源基因质粒