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The PCR products for each gene were purified using Qiagene purification kit. The T7 RNA polymerase gene was digested with NheI and XhoI. Then, after purification with a gel extraction kit (Qiagene), the DNA fragment of T7 RNA polymerase (in length of 2600Kb) was cloned into pIRES2-EGFP plasmid (clontech) and recombinant vector was called pIRES-T7. The cloning process for N and P genes were similar. The PCR products for each gene was purified and digested with NotI. The NotI site designed in 5’-end of reverse primers, but there was not any restriction enzyme site in forward primers. The forward primers contained a kozak consensus ribosme binding site (AACC) and ATG initiation codon. The pIRES2-EGFP plasmid was digested in a step by step process. First, pIRES2-EGFP was digested with BstxI and then, the digestion product of the plasmid treated by klenow to produce blunt end. Finally, pIRES2-EGFP was digested with NotI. The DNA fragments of N and P genes cloned into pIRES2-EGFP and recombinant vectors were called pIRES-N and pIRES-P, respectively. To produce tricistronic expression vecto...

Dead Man’s Shoes †Essay Essay

The short story â€Å"Dead Man’s Shoes† (1998) by David Evans is an astonishing view into the life of a rural village in South Africa after the fall of apartheid, and of the different attitudes flourishing in the community. The story depicts the life of a rich widow, Anne Bezuidenhout, who lives alone on an enormous farm with only her black workers to help her. She has many offers of marriage, all with something to offer her, but she ends up surprising everyone with her choice. However, at the heart of the story we see the persistence of apartheid and conservative values in the ‘new South Africa’. Perhaps one can see this persistence in the narrative voice of the short story. As a 1st person narrator, he is part of the story, as one of the seven suitors of Anne; â€Å"Last and least me, a teacher, divorced by a wife who had found me, my profession and Pampoenfontein too dull for her.† The story is slightly affected by the narrator’s opinions. This is visible in several quotes such as â€Å"Most important, she was rich.† and â€Å"But a good Black staff was one thing. A woman trying to manage it on her own was quite another.† The last quote is one of several quotes in the short story pointing towards the narrators both racist and slightly patriarchal mind. One could argue that the quote just pities Anne for being alone on the farm, however on page 3 Anne tells the suitors that she is in fact used to hardship. If one reads between the lines, it is clear that she is aware of her situation but is not worried about it at all. The language in the story al so has a role in the slightly racist values of the narrator. We see this in the choice of words the author has made when describing, for instance, Anne and Samuel. In the description of Anne, the author uses many positive words to do a detailed image of her, as seen on page 1; â€Å"pleasant-faced with wide green eyes and a voice which carried soft currents of her native Galway. She also had a flickering smile charming and at the same time tantalizing, hinting somehow at private amusement and undeclared opinions†. Naturally Anne would receive a detailed introduction as she is one of the main characters in the story, yet Samuel is also a main character, but his introduction is much less personal and without many adjectives, as seen on page 3: â€Å"Anne had taught him to read and write and to do simple sums. This bit of education hadn’t  spoiled him as it did so many. He knew his place: bossboy among the Black workers†. On top of that the author has let the characters use several nicknames for Samuel such as â€Å"swartg oed† (page 5) and â€Å"boy†(page 4). The use of nicknames could, however, lead the reader on to having a look at the characters of the story. As a last contributor to the general feeling of continued apartheid in the story is the characterization. The characters are revealed through an outward description. In the beginning of the story, the introduction of Anne is given very quickly and detailed. But the author’s way of describing the seven suitors is almost like a list where each of them equals a different personality or genetic trait such as â€Å"Harry Smith, the town’s auctioneer, was the oldest†, â€Å"Japie van Os (†¦) was the richest†, â€Å"Hannes Snyman (†¦) was the biggest† and so forth. The reader must therefore make his or her own conclusions on the character through behavior and talking, because of the author’s use of implicit characterization. For instance the reader could focus on the character Maritz Grootbek and immediately conclude that he is indeed a racist. Why? Because he frequently comments on Samuel. One could argue that this is simply because he admires Anne for having â€Å"a good staff with a reliable Black foreman† (page 3), but his rude behavior and ways of talking states otherwise. An example could be when he arrives at Anne’s house and is shocked when Samuel doesn’t fetch her immediately; â€Å"†Where’s the Nkosikazi (frue), boy?† Jamie demanded roughly. â€Å"Go and fetch her. Hurry!†. Since the other suitor doesn’t argue with his rudeness it is clear that they share his opinions. Last but not least, the shared racist values of the seven suitors become certain when Anne tricks them and decides to marry Samuel. Based on their reaction â€Å"We stared, we glared, we blinked in disbelief, then stared again.† (page 5), it is clear that none of them ever thought that they would be put aside in favor of a black man and even after she has announced her will they still â€Å"tried to dissuade Anne† (page 5). Even though it is generally known between the characters in the story that Samuel is a good man, he is still considered low because of his color, and the suitors are still suspicious about him. Ultimately this shows that even though apartheid has legally ended, it is still a state of mind for many white South Africans, as confirmed by the narrator himself: â€Å"Pampoenfontein may have accepted that all of us – whatever our colour – were equal citizens in the new South Africa, but we  were all proud of our past and no white man there could be expected to tolerate any talk of a black being as good as a white†.

Civil Disobedience is The Cause of Protests - Free Essay Example

Sample details Pages: 2 Words: 540 Downloads: 3 Date added: 2019/05/05 Category Society Essay Level High school Tags: Civil Disobedience Essay Did you like this example? Civil disobedience has been the cause for a large number of protests around the world. It is defined an individuals rights to freedom of speech and is done in a peaceful way. Civil disobedience is important in a democratic country because it allows individuals to exercise their rights and speck against the unfair and unjust government and its laws. Don’t waste time! Our writers will create an original "Civil Disobedience is The Cause of Protests" essay for you Create order Even though the citizens are given their rights, they could be put in jail, could get hurt or even killed for protesting against these laws. It allowed Americans to speak against the Britain and their unjust laws, rules and even against their own government. Gandhi and Thoreau believed that there would be no government without the people it govern, That government is best when it govern least. The government was for the protection of the citizens rights and their opportunities, but instead the government was hurting the people. In these cases the citizens used non violent protest against the laws they thought was unjust and were able to succeed. Henry David Thoreau refused to pay taxes that would fund the American Mexican war, which he was thrown in jail for. He preferred going to prison to get his voice heard without using any violence. Thoreau wanted there to be an end to slavery, the end of the wars in Mexico and an end to unnecessary taxes. Gandhi highly influenced by Thoreau ,was thrown in prison many times because he was willing to not obey the laws to help raise awareness to the people who didnt know what was going on. Even though he was in prison during the Salt march that didnt stop the people from marching on. He didnt give up on what he believed and because of that his non violent march helped India gain freedom from the British. Another man that helped make a b ig impact on the world today by using civil disobedience is Martin Luther King, he used his civil disobedience strategy to also protest against government laws he thought were unjust. This took a large amount of people who were willing to putter life the line so that the laws could be fair. Gandhis ideas influenced King to fight for civil rights peacefully. King was arrested and thrown into jail where he wrote his Letter from Birmingham jail to a clergyman. The purpose of the letter was to address the people who criticized him about what he was doing when he was only doing what was wrong and he let the people who were against what he was doing know that what they are doing and how they were treating the blacks wasnt right. He continued to fight for the rights for blacks. Freedom must be demanded by the oppressed ( MLK). He wanted the protesters to know that if they wanted to gain their freedom and have the same rights everyone else have they have to come together peacefully and let the world know what is going on.These men used civil disobedience to help change unfair laws and to raise awareness about a particular situation. It wasnt used for personal gain, and th eir intentions were to remain peaceful and because they used it for all the right reasons the citizens won their civil rights.