新型冠狀病毒（SARS-CoV-2）引發的新冠肺炎（COVID-19）從2019年12月零號病人確診至今，在全球各地發展為持續的疫情。造成大量患者患病甚至死亡的同時，亦出現了多種變異株。前人的研究中定義了多種病毒株，以及其各自的代表性的突變。然而，在新型冠狀病毒基因組中各片段所遭受的選汰壓力以及各變異株間演化上的異同上，仍缺乏系統性的了解。為更系統性地了解此病毒的演化及變異程度，我分析了2020年1月至2021年8月間美國的55,418條新冠病毒基因組序列。本研究計算新冠病毒中12段蛋白質編碼序列的非同義替換率(non-synonymous substitution rate, dN)和同義替換率（synonymous substitution rate, dS），發現ORF1a、ORF1b、ORF3a、ORF7a, matrix, nucleocapsid和spike的非同義與同義替換率之比例dN/dS ratio有增大的趨勢，顯示出這些基因可能受到正向選擇壓力的影響。雖然dN/dS分析幫助我找出受正向選擇的基因，但是無法找出受正向選擇的位點。因此，為找出有較大機會受到正向選擇影響的位點，我找出在各變異株中隨時間顯著增加的突變，並在spike蛋白質結構中標示出這些突變的位置。透過上述的分析，我總共找出了100個較有機會發生重要突變的位點，共125種突變。其中包含了過去曾在其他研究中被提及的位點，以及尚未被注意到的位點。我的研究，對新冠病毒的演化，尤其是自然選汰，提供了全方面的探討與見解。

The COVID-19 caused by SARS-CoV-2 has developed into a global pandemic which leads to about four hundred million cases and over five million deaths worldwide since the first case reported in December 2019. Several strains of SARS-CoV-2 have been identified by mutations in previous studies. However, a comprehensive understanding of selective pressure effect on each protein of SARS-CoV-2 is still lacking. In this study, I analyzed 55,418 SARS-CoV-2 genome sequences collected from the United States between January 2020 and August 2021. I calculated the dN and dS of 12 protein-coding sequences. There is an increasing tendency of dN/dS in ORF1a, ORF1b, ORF3a, ORF7a, matrix, nucleocapsid and spike, indicating that these proteins may affected by positive selection. To found sites with a greater chance of being affected by positive selection, I identified 125 mutations happened on 100 site that increased significantly over time in each variant and colored these sites in the spike protein. Our research provides comprehensive discussions and insights into the evolution and selection of SARS-CoV-2.

摘要 ii
Abstract iii
序言 iv
目錄 vi
圖目錄 vii
表目錄 viii
第一章 背景介紹 1
第二章 材料與方法 6
2.1序列資料的獲得與處理 6
2.2分派Nextstrain clade，clade內演化分析 7
2.3計算dN、dS 8
第三章 結果與討論 10
3.1變異株的改變 10
3.2自然選汰分析 10
3.3位點分析 13
第四章 總結 15
第五章 圖片與表格 16
第六章 參考文獻 37

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