South Dakota State University Open PRAIRIE: Open Public Research Access Institutional Repository and Information Exchange Theses and Dissertations 2017 Investigation of Candidate Loci Associated with Maize Perennialism Anjun Ma South Dakota State University Follow this and additional works at: http://openprairie.sdstate.edu/etd Part of the Agriculture Commons, and the Plant Biology Commons Recommended Citation Ma, Anjun, "Investigation of Candidate Loci Associated with Maize Perennialism" (2017). Theses and Dissertations. 1701. http://openprairie.sdstate.edu/etd/1701 This Thesis - Open Access is brought to you for free and open access by Open PRAIRIE: Open Public Research Access Institutional Repository and Information Exchange. It has been accepted for inclusion in Theses and Dissertations by an authorized administrator of Open PRAIRIE: Open Public Research Access Institutional Repository and Information Exchange. For more information, please contact [email protected]. INVESTIGATION OF CANDIDATE LOCI ASSOCIATED WITH MAIZE PERENNIALISM BY ANJUN MA A thesis submitted in partial fulfillment of the requirements for the Master of Science Major in Biological Sciences South Dakota State University 2017 iii ACKNOWLEDGEMENTS I would like to express my gratitude to my advisor Dr. Yang Yen for his guidance and care in the past three years. With his full help, I can eventually complete my research and step forward. I also want to thank my committee members: Dr. Yajun Wu, Dr. Qin Ma and Dr. Cristopher Schmit for their help in the design and implementation of my experiments, data analysis and manuscript preparation. I would also like to thank Dr. Donald Auger, Dr. Jixiang Wu and Dr. Frank You for their kindly advises and comments to my research. My sincere thankfulness goes beyond to my lab mates Yinjie Qiu, Dr. Yongbin Zhuang, Subha Dahal, Bimal Paudel and Tajbir Raihan for their help with the lab work and helpful discussion. All my friends in SDSU: Zhao Wang, Jieshi Yu, Yiran Zhang and many others, supported me not only in work but also in life. It is my honor to have them. Most importantly, I would like to thank my beloved parents. Without their encouragement and support I can never be successful. iv CONTENT ACKNOWLEDGEMENTS ............................................................................................... iii CONTENT ......................................................................................................................... iv LIST OF FIGURES ........................................................................................................... vi LIST OF TABLES ............................................................................................................ vii ABBREVIATIONS ......................................................................................................... viii ABSTRACT .........................................................................................................................x CHAPTER 1. LITERATURE REVIEW .............................................................................1 1.1 Overview of crop production and issues caused by agriculture .................................1 1.2 The advantage of perennial plants ..............................................................................2 1.3 Progress of perennial crops ........................................................................................2 1.4 Genotyping-by-sequencing.........................................................................................5 References ........................................................................................................................8 CHAPTER 2 INVESTIGATION OF CANDIDATE LOCI ASSOCIATED WITH MAIZE PERENNIALISM.................................................................................................16 2.1 Introduction ..............................................................................................................16 2.2 Materials and methods ..............................................................................................20 2.2.1 Population generation ........................................................................................ 20 2.2.2 DNA preparation ............................................................................................... 21 2.2.3 Genotype-by-sequencing ................................................................................... 21 2.2.4 TASSEL pipeline and SNP discovery ............................................................... 22 2.2.5 Scoring genotypes and phenotypes ................................................................... 23 2.2.6 Missing data fixing and chi-square test ............................................................. 23 2.2.7 Locus test ........................................................................................................... 25 2.2.8 Verification of the predicted loci....................................................................... 26 2.3 Results ......................................................................................................................28 2.3.1 Phenotypic data ................................................................................................. 28 2.3.2 SNP discovery and genotypic data .................................................................... 29 2.3.3 Chi-square goodnees of fit test and missing data fixation................................. 30 2.3.4 Analysis of physical intervals............................................................................ 30 v 2.3.5 Verification of the predicted loci....................................................................... 32 2.4 Discussion.................................................................................................................33 2.4.1 Determination of perennialism .......................................................................... 33 2.4.2 Chromosome 2 and 7 might be associated with perenniality in maize ............. 34 2.4.3 SNP Marker genotyping indicated limited correlation...................................... 35 2.5 Conclusion and future perspectives ..........................................................................36 References ......................................................................................................................38 Appendix ............................................................................................................................58 Appendix 1. Steps in TASSEL pipeline including example command lines and brief descriptions ...............................................................................................................58 Appendix 2A.R codes for candidate locus analysis .......................................................61 Appendix 2B.R code for finding aligned sequences ......................................................62 Appendix 3. Chi-Square goodness of fit test and LOD score for all 621 significant SNPs after the 4th filter .......................................................................................................63 Appendix 4.All tested primers for selected position. .....................................................77 vi LIST OF FIGURES Figure 1. 1 Comparison of root systems of annual and perennial wheat .......................... 14 Figure 2. 1 The general process of TASSEL pipeline ......................................................42 Figure 2. 2 Regrowth phenotype shown in BZ F1 ............................................................43 Figure 2. 3 Different forms of non-regrowth and regrowth ..............................................44 Figure 2. 4 Regrowth appear at knots and at basal ...........................................................45 Figure 2. 5 Genetic map of SNP markers distribution across 10 chromosomes. ..............46 Figure 2. 6 LOD scores for candidate locus determination ..............................................47 Figure 2. 7 Effect plots for two candidate positions .........................................................48 Figure 2. 8 Genotyping of 2-1A primer set.......................................................................49 vii LIST OF TABLES Table 1. 1 Top agricultural production by individual crops in 2014 ................................15 Table 2. 1 Barcodes for all 95 individuals ........................................................................50 Table 2. 2 SNP numbers in each chromosome after every filter step. ..............................51 Table 2. 3 BZ F2 Phenotype scores ...................................................................................52 Table 2. 4 94 samples and Z. diploperennis used for GBS with phenotypic scores. ........53 Table 2. 5 Candidate genes for SNP. ................................................................................54 Table 2. 6 SNP primers designed for PCR mapping (polymorphic) ................................55 Table 2. 7 The contiguous table for χ2 test of 2-1A mapping result .................................56 Table 2. 8 Genes found overlapped or were close to 2-1A SNP ......................................57 viii ABBREVIATIONS AS-PCR Allele-specific polymerase chain reaction bp Base pair Chr Chromosome CTAB Cetyl trimethylammonium bromide DNA Deoxyribonucleic acid GBS Genotype-by-sequencing gt1 grassy tiller 1 id1 Indeterminate 1 KASP Kompetitive Allele Specified PCR LOD Logarithm of odds MAF Minimum allele frequency ng Nano gram