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.
summary
Bacterial Resistance(s): Ampicillin
Growth Temperature: 37°C
Growth Strain(s): TG1
Copy number: High Copy
Insert Size (bp): 1600
Promoter:lacZ
Tags / Fusion Proteins
HA tag (C terminal on backbone)
6x His (C terminal on backbone)
geneIII (C terminal on backbone)
5′ sequencing primer
5'-AAG ACA GCT ATC GCG ATT GCA G-3'
3′ sequencing primer
5'-GCC CCC TTA TTA GCG TTT GCC ATC-3'
Vector sequence
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
//
