I have to produce a vaccine using recombinant DNA technology. The gene i want to amplify is to be expressed in a Yeast vector. But i am suppose to DESIGN 2 sets of primers! One without his tag and the other with his tag. I will amplify the gene, subclone it in pGEMT, the clone in pET28a(+) which come with his tag. Purify and finally clone in a yeast vector (pKOV96) and express in yeast cells.
How do i go about designing both sets of primers??????? I have to amplify the gene from the N- to the C-terminal to increase the immunogenecity of my vaccine. Can anyone please help? Here is the Clostridium perfringens strain DNA:
ATGAAGAAAAAATTTATTTCATTAGTTATAGTTAGTTCACTTTTAAACGGATGCCTATTATCACCAACTT
TAGTGTATGCAAATGATATAGGTAAAACTACTACTATAACTAGAAATAAGACATCAGATGGCTATACTAT
AATTACACAAAATGATAAACAGATAATATCATATCAATCTGTTGACTCTTCAAGTAAAAATGAAGATGGT
TTTACTGCATCTATAGATGCTAGATTTATCGATGATAAATATTCATCTGAAATGACAACTTTAATAAACT
TAACTGGATTTATGTCTTCAAAAAAAGAAGATGTTATAAAAAAATACAATTTGCATGATGTTACTAATTC
TACTGCAATTAATTTTCCGGTTAGATACTCGATTTCTATTTTAAATGAAAGTATTAATGAAAATGTAAAA
ATAGTTGATAGTATTCCTAAAAATACAATTTCTCAAAAAACTGTATCCAATACAATGGGATACAAAATAG
GAGGTTCAATTGAAATAGAAGAAAATAAACCTAAAGCTTCAATTGAAAGCGAATATGCTGAATCATCTAC
AATAGAATATGTCCAACCTGATTTTTCTACTATACAGACAGATCATTCAACCTCTAAAGCTTCATGGGAT
ACAAAATTTACAGAAACTACTCGTGGTAATTATAATTTAAAATCAAACAACCCTGTATATGGAAATGAAA
TGTTTATGTACGGAAGATATACTAATGTTCCTGCAACTGAAAATATAATTCCAGATTATCAAATGTCAAA
ATTAATAACAGGTGGTTTAAACCCTAATATGTCTGTAGTTCTAACTGCTCCTAATGGTACTGAAGAATCT
ATAATAAAAGTTAAAATGGAGCGTGAAAGAAACTGTTATTATCTTAATTGGAATGGTGCTAACTGGGTAG
GACAAGTCTATTCCAGGCTAGCTTTTGATACCCCAAATGTAGATAGTCATATATTTACATTCAAAATAAA
TTGGCTTACTCACAAAGTAACAGCTATTTAG