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. The “TT” refers to the human Fab to tetanus toxin which can be used as a template to amplify the kappa light chain constant region and Fd for construction of chimeric antibody libraries .
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
6x His (C terminal on backbone)
HA tag (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 ctcacgcagt
301 ctccaggcac cctgtctttg tctccagggg aaagagccac cctctcctgc agggccagtc
361 acagtgttag cagggcctac ttagcctggt accagcagaa acctggccag gctcccaggc
421 tcctcatcta tggtacatcc agcagggcca ctggcatccc agacaggttc agtggcagtg
481 ggtctgggac agacttcact ctcaccatca gcagactgga gcctgaagat tttgcagtgt
541 actactgtca gcagtatggt ggctcaccgt ggttcggcca agggaccaag gtggaactca
601 aacgaactgt ggctgcacca tctgtcttca tcttcccgcc atctgatgag cagttgaaat
661 ctggaactgc ctctgttgtg tgcctgctga ataacttcta tcccagagag gccaaagtac
721 agtggaaggt ggataacgcc ctccaatcgg gtaactccca ggagagtgtc acagagcagg
781 acagcaagga cagcacctac agcctcagca gcaccctgac gctgagcaaa gcagactacg
841 agaaacacaa agtctacgcc tgcgaagtca cccatcaggg cctgagcttg cccgtcacaa
901 agagcttcaa caggggagag tgttagttct agataattaa ttaggaggaa tttaaaatga
961 aatacctatt gcctacggca gccgctggat tgttattact cgctgcccaa ccagccatgg
1021 ccgaggtgca gctgctcgag cagtctgggg ctgaggtgaa gaagcctggg tcctcggtga
1081 aggtctcctg cagggcttct ggaggcacct tcaacaatta tgccatcagc tgggtgcgac
1141 aggcccctgg acaagggctt gagtggatgg gagggatctt ccctttccgt aatacagcaa
1201 agtacgcaca acacttccag ggcagagtca ccattaccgc ggacgaatcc acgggcacag
1261 cctacatgga gctgagcagc ctgagatctg aggacacggc catatattat tgtgcgagag
1321 gggatacgat ttttggagtg accatgggat actacgctat ggacgtctgg ggccaaggga
1381 ccacggtcac cgtctccgca gcctccacca agggcccatc ggtcttcccc ctggcaccct
1441 cctccaagag cacctctggg ggcacagcgg ccctgggctg cctggtcaag gactacttcc
1501 ccgaaccggt gacggtgtcg tggaactcag gcgccctgac cagcggcgtg cacaccttcc
1561 cggctgtcct acagtcctca ggactctact ccctcagcag cgtggtgacc gtgccctcca
1621 gcagcttggg cacccagacc tacatctgca acgtgaatca caagcccagc aacaccaagg
1681 tggacaagaa agttgagccc aaatcttgtg acaaaactag tggccaggcc ggccagcacc
1741 atcaccatca ccatggcgca tacccgtacg acgttccgga ctacgcttct taggagggtg
1801 gtggctctga gggtggcggt tctgagggtg gcggctctga gggaggcggt tccggtggtg
1861 gctctggttc cggtgatttt gattatgaaa agatggcaaa cgctaataag ggggctatga
1921 ccgaaaatgc cgatgaaaac gtgctacagt ctgacgctaa aggcaaactt gattctgtcg
1981 ctactgatta cggtgctgct atcgatggtt tcattggtga cgtttccggc cttgctaatg
2041 gtaatggtgc tactggtgat tttgctggct ctaattccca aatggctcaa gtcggtgacg
2101 gtgataattc acctttaatg aataatttcc gtcaatattt accttccctc cctcaatcgg
2161 ttgaatgtcg cccttttgtc tttggcgctg gtaaaccata tgaattttct attgattgtg
2221 acaaaataaa cttattccgt ggtgtctttg cgtttctttt atatgttgcc acctttatgt
2281 atgtattttc tacgtttgct aacatactgc gtaataagga gtcttaagct agctaattaa
2341 tttaagcggc cgcagatctg ctctctgagg aggatctggg aaattgtaag cgttaatatt
2401 ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat tttttaacca ataggccgaa
2461 atcggcaaaa tcccttataa atcaaaagaa tagaccgaga tagggttgag tgttgttcca
2521 gtttggaaca agagtccact attaaagaac gtggactcca acgtcaaagg gcgaaaaacc
2581 gtctatcagg gcgatggccc actacgtgaa ccatcaccct aatcaagttt tttggggtcg
2641 aggtgccgta aagcactaaa tcggaaccct aaagggagcc cccgatttag agcttgacgg
2701 ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag cgaaaggagc gggcgctagg
2761 gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca cacccgccgc gcttaatgcg
2821 ccgctacagg gcgcgtcagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt
2881 ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg
2941 cttcaataat attgaaaaag gaagagtatg agtattcaac atttccgtgt cgcccttatt
3001 cccttttttg cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta
3061 aaagatgctg aagatcagtt gggtgcacga gtgggttaca tcgaactgga tctcaacagc
3121 ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc caatgatgag cacttttaaa
3181 gttctgctat gtggcgcggt attatcccgt attgacgccg ggcaagagca actcggtcgc
3241 cgcatacact attctcagaa tgacttggtt gagtactcac cagtcacaga aaagcatctt
3301 acggatggca tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact
3361 gcggccaact tacttctgac aacgatcgga ggaccgaagg agctaaccgc ttttttgcac
3421 aacatggggg atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata
3481 ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta
3541 ttaactggcg aactacttac tctagcttcc cggcaacaat taatagactg gatggaggcg
3601 gataaagttg caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat
3661 aaatctggag ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt
3721 aagccctccc gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga
3781 aatagacaga tcgctgagat aggtgcctca ctgattaagc attggtaact gtcagaccaa
3841 gtttactcat atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag
3901 gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac
3961 tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc
4021 gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat
4081 caagagctac caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat
4141 actgtccttc tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct
4201 acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt
4261 cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg
4321 gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta
4381 cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg
4441 gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg
4501 tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc
4561 tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg
4621 gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat
4681 aaccgtatta ccgcctttga gtgagctgat accgctcgcc gcagccgaac gaccgagcgc
4741 agcgagtcag tgagcgagga agcgga
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