pComb3X SS

$215.00

  • Model: PVT10572
  • 48 Units in Stock
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pComb3X SS


Catalog No. PVT10572
Packing 2ug

 

pComb3X SS Description

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.

 

pComb3X SS Information

Promoter: Lac/lac promoter

Plasmid size: 4991 BP

Plasmid Label: C-6*HIS, C-HA

Prokaryotic resistance: ampicillin Amp

Cloning strain: Escherichia coli Stbl3

Culture conditions: 37 C, aerobic, LB

 

pComb3X SS Sequence

LOCUS       Exported                4991 bp ds-DNA     circular SYN 27-OCT-2017

DEFINITION  synthetic circular DNA

SOURCE      synthetic DNA construct

  ORGANISM  synthetic DNA construct

REFERENCE   1  (bases 1 to 4991)

FEATURES             Location/Qualifiers

     source          1..4991

                     /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)."

     misc_feature    294..1470

                     /label=Light Chain

     CDS             complement(574..591)

                     /codon_start=1

                     /product="6xHis affinity tag"

                     /label=6xHis

                     /translation="HHHHHH"

     misc_feature    1500..1577

                     /label=PelB Leader

     CDS             1584..1907

                     /codon_start=1

                     /gene="trxA"

                     /product="E. coli thioredoxin"

                     /label=Heavy Chain

                     /translation="MSDKIIHLTDDSFDTDVLKADGAILVDFWAEWCGPCKMIAPILDE

                     IADEYQGKLTVAKLNIDQNPGTAPKYGIRGIPTLLLFKNGEVAATKVGALSKGQLKEFL

                     DANL"

     CDS             1962..1979

                     /codon_start=1

                     /product="6xHis affinity tag"

                     /label=6xHis

                     /translation="HHHHHH"

     CDS             1986..2012

                     /codon_start=1

                     /product="HA (human influenza hemagglutinin) epitope tag"

                     /label=HA

                     /translation="YPYDVPDYA"

     CDS             2118..2552

                     /codon_start=1

                     /label=geneIII

                     /translation="MANANKGAMTENADENVLQSDAKGKLDSVATDYGAAIDGFIGDVS

                     GLANGNGATGDFAGSNSQMAQVGDGDNSPLMNNFRQYLPSLPQSVECRPFVFGAGKPYE

                     FSIDCDKINLFRGVFAFLLYVATFMYVFSTFANILRNKES"

     rep_origin      complement(2606..3061)

                     /direction=LEFT

                     /label=f1 ori

                     /note="f1 bacteriophage origin of replication; arrow 

                     indicates direction of (+) strand synthesis"

     promoter        3088..3192

                     /gene="bla"

                     /label=AmpR promoter

     CDS             3193..4053

                     /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      4224..4812

                     /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 ctcgccatgg

      301 ctggttgggc agcgagtaat aacaatccag cggctgccgt aggcaatagg tatttcatta

      361 tgactgtctc cttggcgact agctagttta gaattcgtaa tcatggtcat agctgtttcc

      421 tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa gcataaagtg

      481 taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc gctcactgcc

      541 cgctttccag tcgggaaacc tgtcgtgtta ctaatgatgg tgatggtgat ggctagtttt

      601 gtcacaagat ttgggctcaa ctttcttgtc caccttggtg ttgctgggct tgtgattcac

      661 gttgcagatg taggtctggg tgcccaagct gctggagggc acggtcacca cgctgctgag

      721 ggagtagagt cctgaggact gtaggacagc cgggaaggtg tgcacgccgc tggtcagggc

      781 gcctgagttc cacgacaccg tcgccggttc ggggaagtag tccttgacca ggcagcccag

      841 ggccgctgtg cccccagagg tgctcttgga ggagggtgcc agggggaaga ccgatgggcc

      901 cttggtggag gctgcggaga cggtgaccgt ggtaccagca gaaacctggc caggctccca

      961 ggctcctcat ctatggtaca tccagcaggg ccactggcat cccagacagg ttcagtggca

     1021 gtgggtctgg gacagacttc actctcacca tcagcagact ggagcctgaa gattttgcag

     1081 tgtactactg tcagcagtat ggtggctcac cgtggttcgg ccaagggacc aaggtggaac

     1141 tcaaacgaac tgtggctgca ccatctgtct tcatcttccc gccatctgat gagcagttga

     1201 aatctggaac tgcctctgtt gtgtgcctgc tgaataactt ctatcccaga gaggccaaag

     1261 tacagtggaa ggtggataac gccctccaat cgggtaactc ccaggagagt gtcacagagc

     1321 aggacagcaa ggacagcacc tacagcctca gcagcaccct gacgctgagc aaagcagact

     1381 acgagaaaca caaagtctac gcctgcgaag tcacccatca gggcctgagc ttgcccgtca

     1441 caaagagctt caacagggga gagtgttagt tctagataat taattaggag gaatttaaaa

     1501 tgaaatacct attgcctacg gcagccgctg gattgttatt actcgctgcc caaccagcca

     1561 tggccgaggt gcagctgctc gagatgagcg ataaaattat tcacctgact gacgacagtt

     1621 ttgacacgga tgtactcaaa gcggacgggg cgatcctcgt cgatttctgg gcagagtggt

     1681 gcggtccgtg caaaatgatc gccccgattc tggatgaaat cgctgacgaa tatcagggca

     1741 aactgaccgt tgcaaaactg aacatcgatc aaaaccctgg cactgcgccg aaatatggca

     1801 tccgtggtat cccgactctg ctgctgttca aaaacggtga agtggcggca accaaagtgg

     1861 gtgcactgtc taaaggtcag ttgaaagagt tcctcgacgc taacctggcg tacccgtacg

     1921 acgttccgga ctacggttct actagtggcc aggccggcca gcaccatcac catcaccatg

     1981 gcgcataccc gtacgacgtt ccggactacg cttcttagga gggtggtggc tctgagggtg

     2041 gcggttctga gggtggcggc tctgagggag gcggttccgg tggtggctct ggttccggtg

     2101 attttgatta tgaaaagatg gcaaacgcta ataagggggc tatgaccgaa aatgccgatg

     2161 aaaacgtgct acagtctgac gctaaaggca aacttgattc tgtcgctact gattacggtg

     2221 ctgctatcga tggtttcatt ggtgacgttt ccggccttgc taatggtaat ggtgctactg

     2281 gtgattttgc tggctctaat tcccaaatgg ctcaagtcgg tgacggtgat aattcacctt

     2341 taatgaataa tttccgtcaa tatttacctt ccctccctca atcggttgaa tgtcgccctt

     2401 ttgtctttgg cgctggtaaa ccatatgaat tttctattga ttgtgacaaa ataaacttat

     2461 tccgtggtgt ctttgcgttt cttttatatg ttgccacctt tatgtatgta ttttctacgt

     2521 ttgctaacat actgcgtaat aaggagtctt aagctagcta attaatttaa gcggccgcag

     2581 atctgctctc tgaggaggat ctgggaaatt gtaagcgtta atattttgtt aaaattcgcg

     2641 ttaaattttt gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg caaaatccct

     2701 tataaatcaa aagaatagac cgagataggg ttgagtgttg ttccagtttg gaacaagagt

     2761 ccactattaa agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat

     2821 ggcccactac gtgaaccatc accctaatca agttttttgg ggtcgaggtg ccgtaaagca

     2881 ctaaatcgga accctaaagg gagcccccga tttagagctt gacggggaaa gccggcgaac

     2941 gtggcgagaa aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta

     3001 gcggtcacgc tgcgcgtaac caccacaccc gccgcgctta atgcgccgct acagggcgcg

     3061 tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata

     3121 cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga

     3181 aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca

     3241 ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat

     3301 cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag

     3361 agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc

     3421 gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct

     3481 cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca

     3541 gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt

     3601 ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat

     3661 gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt

     3721 gacaccacga tgcctgtagc aatggcaaca acgttgcgca aactattaac tggcgaacta

     3781 cttactctag cttcccggca acaattaata gactggatgg aggcggataa agttgcagga

     3841 ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt

     3901 gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc

     3961 gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct

     4021 gagataggtg cctcactgat taagcattgg taactgtcag accaagttta ctcatatata

     4081 ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt

     4141 gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc

     4201 gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg

     4261 caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact

     4321 ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt ccttctagtg

     4381 tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg

     4441 ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac

     4501 tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca

     4561 cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga

     4621 gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc

     4681 ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct

     4741 gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg

     4801 agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct

     4861 tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc

     4921 tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc

     4981 gaggaagcgg a

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