Fusion immunotoxin ML-L-SEC2 and gene and its preparation
An ML-L-SEC2, immunotoxin technology, applied in the field of genetic engineering pharmacy, can solve the problems of side effects, lack of specificity in tumor treatment, and achieve the effect of eliminating side effects
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Embodiment 1
[0024] A humanized anti-HER-2 single-chain antibody gene ml3.9 has the base sequence of SEQ ID No: 3 in the sequence table.
[0025] ATGGCCCAGG TGCAGCTGGT GCAGTCTGGG GCAGAGGTGA AAAAGCCCGG 50
[0026] GGAGTCTCTG AAGATCTCCT GTAAGGGTTC TGGATACAGC TTTACCAGCT 100
[0027] ACTGGATCGC CTGGGTGCGC CAGATGCCCG GGAAAGGCCT GGAGTACATG 150
[0028] GGGCTCATCT ATCCTGGTGA CTCTGACACC AAATACAGCC CGTCCTTCCA 200
[0029] AGGCCAGGTC ACCATCTCAG TCGACAAGTC CGTCAGCACT GCCTACTTGC 250
[0030] AATGGAGCAG TCTGAAGCCC TCGGACAGCG CCGTGTATTT TTGTGCGAGA 300
[0031] CATGACGTGG GATATTGCAG TAGTTCCAAC TGCGCAAAGT GGCCTGAATA 350
[0032] CTTCCAGCAT TGGGGCCAGG GCACCCTGGT CACCGTCTCC TCAGGTGGAG 400
[0033] GCGGTTCAGG CGGAGGTGGC TCTGGCGGTG GCGGATCGCA GTCTGTGTTG 450
[0034] ACGCAGCCGC CCTCAGTGTC TGCGGCCCCA GGACAGAAGG TCACCATCTC 500
[0035] CTGCTCTGGA AGCAGCTCCA ACATTGGGAA TAATTATGTA TCCTGGTACC 550
[0036] AGCAGCTCCC AGGAACAGCC CCCAAACTCC TCATCTATGA TCACACCAAT 600
[0037] CGGCCCGCAG GGGTCCCTGA CCGATTCTCT G...
Embodiment 2
[0062] Staphylococcus aureus enterotoxin C2 (SEC2) gene sec2 is derived from Staphylococcus aureus and has the base sequence of SEQ ID No: 5 in the sequence table.
[0063] GAGAGTCAACCAGACCCTACGCCAGATGAGTTGCACAAATCAAGTGAGTTTACTGGTACGATGGGTAATATGAAATATTTATATGATGATCATTATGTATCAGCAACTAAAGTTATGTCTGTAGATAAATTTTTGGCACATGATTTAATTTATAACATTAGTGATAAAAAACTAAAAAATTATGACAAAGTGAAAACAGAGTTATTAAATGAAGATTTAGCAAAGAAGTACAAAGATGAAGTAGTTGATGTGTATGGATCAAATTACTATGTAAACTGCTATTTTTCATCCAAAGATAATGTAGGTAAAGTTACAGGTGGTAAAACTTGTATGTATGGAGGAATAACAAAACATGAAGGAAACCACTTTGATAATGGGAACTTACAAAATGTACTTATAAGAGTTTATGAAAATAAAAGAAACACAATTTCTTTTGAAGTGCAAACTGATAAGAAAAGTGTAACAGCTCAAGAACTAGACATAAAAGCTAGGAATTTTTTAATTAATAAAAAAAATTTGTATGAGTTTAACAGTTCACCATATGAAACAGGATATATAAAATTTATTGAAAATAACGGCAATACTTTTTGGTATGATATGATGCCTGCACCAGGCGATAAGTTTGACCAATCTAAATATTTAATGATGTACAACGACAATAAAACGGTTGATTCTAAAAGTGTGAAGATAGAAGTCCACCTTACAACAAAGAATGGA
[0064] (1) Information of SEQ ID No: 5 (see sequence listing)
[0065] (A) Sequential features
[...
Embodiment 3
[0086] The DNA Linker sequence encoding the connecting short peptide has the following base sequence (oligonucleotide chain):
[0087] GGCCGCAGGTGGAGGCGGTTCAGGCGGAGGTGGCTCTGAATTC
[0088] (1) Preparation of DNA Linker sequence gene
[0089] 1) Design and synthesis of DNA Linker sequence gene:
[0090] Design and synthesize two oligonucleotide strands:
[0091] Forward strand L-F: 5'- G GCC GC A AGC GGC TCA GGA TCT GGA TCA GGA TCTGGC G-3';
[0092] Reverse strand L-R: 5'- AAA TTC GCC AGA TCC TGA TCC AGA TCC TGA GCCGCT TGC-3'
[0093] Add NotI and EcoRI sticky ends to the upstream and downstream respectively, and the encoded glycine serine short peptide is (GS) 2 sequence.
[0094] 2) Annealing of oligonucleotide chains:
[0095] 10×Annealing Hybridization Buffer (SSC) Components: 1.5mol / L NaCl, 0.3M Trisodium Citrate·2H 2 O, adjust the pH to 7.0 with HCl;
[0096] Annealing reaction system: 10×SSC buffer 1μl, positive and negative oligonucleotide chains 5pmol ...
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