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研究生:卓冠佑
研究生(外文):CHO, KUAN-YU
論文名稱:提高水稻T-DNA兩翼序列解序成功率
論文名稱(外文):Research of Improving Success Ratio of T-DNA Flanking Sequence Protocol
指導教授:范宗宸
指導教授(外文):FAN, MING-JEN
口試委員:鍾景光范宗宸黃元勵
口試委員(外文):CHUNG, JING-GUNGFAN, MING-JENHUANG, YUAN-LI
口試日期:2017-07-21
學位類別:碩士
校院名稱:亞洲大學
系所名稱:生物科技學系
學門:生命科學學門
學類:生物科技學類
論文種類:學術論文
論文出版年:2017
畢業學年度:105
語文別:中文
論文頁數:100
中文關鍵詞:兩翼序列定序
外文關鍵詞:Flanking sequenceT-DNAPCR
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水稻rice (Oryza sativa L.),禾本科單子葉植物,為重要的糧食作物之一,研究水稻功能性基因主要以T-DNA作為Tag,建立T-DNA插入突變點之 FST資料庫,可加速水稻功能性基因體的研究。中央研究院余淑美所帶領的團隊利用TN67 T-DNA插入突變株建立台灣水稻插入突變體族群及相關資料庫(TRIM),此突變族群及資料庫之建立,對於提供水稻功能性基因體相關研究具極大的幫助 ,而目前建立的TN67 T-DNA插入突變族群,其總數超過120,000個突變植株品系,並委託亞洲大學水稻T-DNA生技研發中心(TTRSC)辦理有關水稻T-DNA插入突變種子之保存及T-DNA插入點兩翼序列之解序。2007年至今,TTRSC的T-DNA種原庫以保存了73,139個水稻T-DNA插入突變品系,並萃取水稻DNA,目前已成功萃取了67,525個T-DNA插入品系的DNA母液,以備進行T-DNA插入點兩翼序列之解序。
TTRSC在2007年開發出的Hooker adaptor PCR方法依照電泳圖會出現的幾種結果分成五大類,其中能夠送解序之成功產物約為54%,再將PCR產物成功之結果進行解序。我們根據利用Hooker5引子的PCR結果的結果分析數據,成功率為53%。總共27878個品系,成功14774解序成功,尚未達到理想,多年來,T-DNA解序是以及左邊界和右邊界之間的DNA序列為Tag,即T-DNA,在右邊界處的T-DNA轉移起始和在左邊界處的終止,有科學家提出了二元載體“骨架”序列可以整合入轉基因植物的DNA中,本研究證實,Backbone序列是解序失敗的主要因素之一,有12%為Backbone,在定序之前將預期結果為Backbone之品系剔除可以降低成本並提高實驗的成功率。
這些資料庫除了可供國內外的學者使用,我們也將這些資料按照T-DNA插入點之排序,從水稻T-DNA生技研發中心所解序的資料庫中篩選出品系M121741、M9944、M122240、M122377、M126390 之FST進行重複序列及SNP的分析。 結果第11條及第12條染色體的插入點上具有高相似度的重複序列,可證明T-DNA可插入在第11條或第12條兩條染色體上,並以NCBI資料庫分析鄰近基因,由結果得知第11條染色體上的插入點含有一個基因(Os11g0143600),將此插入點與此基因進行SNP的分析,結果在插入點與基因之間有C/T、G/C、G/A、G/C、G/A 、A/G、T/C共7個SNP,這些SNP對此基因和性狀表達具有差異性及此基因對插入點之影響,但實際於田間之表達需再進一步探討。
本實驗由國際稻米研究中心(IRRI)委託20個難解之DNA品系進行提升成功率之成果,共18個品系解序成功,成功率提升至80%,而解序成本從原來28600降低至17400,約降低了40%之解序成本。

Rice rice (Oryza sativa L.). Gramineae monocotyledons. As one of the important food. Recent studies on rice functional genes are mainly focus on Transfer-DNA (T-DNA) as a tag. To obtain T-DNA insertion mutations site, Flanking Sequence Tag (FST) is indispensable for reverse genetics studies. Therefore, to stablish T-DNA insertion mutations point of FST database, can accelerate rice functional genomics researches. The establishment of this database for rice functional gene research has a great help. At present the total number of more than 120,000 mutant plant lines. And entrusted the Asian University T-DNA Taged Rice Service Center(TTRSC) to deal with the rice T-DNA insert mutant seed preservation and sequencing flanking sequence of T-DNA insertion site .
The team led by Yu Shu-mei of the Central Research Institute used the TN67 T-DNA insertion mutant to establish the Taiwan rice insert mutant population and related databases(Taiwan Rice Insertional Mutants Database,TRIM).
For many years, it was believed that only the DNA between the repeats, the T-DNA, and not the external vector DNA is transferred to the plant cell. However, recent and more detailed characterization of the DNA inserts in transgenic plants demonstrate that very frequently also vector backbone sequences are integrated into the plant genome.
Sequencing of the T-DNA flanking sequence need to face uncertainties in the PCR experiment and sequencing then blast rice genome and plasmid sequence database. Researcher can confirm the success of the FST in the way which is time-consuming and costly. The data used for this study is Taiwan Rice Insertional Mutants Database. We analysis data according to the results of PCR outcomes to explore the reasons for success and failure and comparing of the superior of PCR method. The successful rate of sequencing is 52%. A total of 27878 lines but 14774 lines is successfully sequenced. It doesn’t achieve the ideal situation. The determinants of failure include plasmid pollution and Backbone sequence. This study has confirmed that the Backbone sequence is one major determinants for the failure of the sequencing. Removing samples of Backbone before sequencing can cut costs and increase the successful rate of the experiment.
This study aimed to use Taiwan rice insertional Mutagens (TRIM) to compare the Flanking Sequence inserted sequence which is found has duplication and compare the single nucleotide polymorphism (SNP) of both site. In this way, the different function of gene insertion site on rice genome can be compared.
This researcher use Basic Local Alignment Search Tool (BLAST) combined Perl programming language to construction a program named the "rice T-DNA insertion mutants Flanking Sequence alignment automation system”. And use this system to distinguish the sequences obtain from PCR, such as of rice T-DNA sequence, plasmid sequence and rice gene sequences. This database is the foundation of rice FST laboratory backboned for analysis FST and BLAST sequence alignment. The research expected to find the same expectations with different insertion point chromosomes. In order to explore the effects of gene (Os11g0143600) insertion points on rice, this study will compare 200bp insertion point sequence to find high similarity of repeats and compare their differences. To analysis the insertion point and gene SNP, the results between the insertion point and a total of seven gene SNP C/T、G/C、G/A、G/C 、G/A 、A/G、T/C. Differential gene expression of a type known these SNP regard, need to do more field research.
In this study, 20 DNA lines were subordinated by IRRI, A total of 18 lines of success order, the success rate increased to 80%, while the replacement cost from the original 28600 down to 17400, about 40% reduction in the cost of the order.

目錄
致謝.............................................................................................................................................................i
摘要.............................................................................................................................................................ii
Abstract......................................................................................................................................................iv
目錄............................................................................................................................................................vi
圖目錄.......................................................................................................................................................viii
表目錄.........................................................................................................................................................x
Abbreviation...............................................................................................................................................xi
第一章 前言................................................................................................................................................1
第二章 前人研究.........................................................................................................................................3
第一節 研究水稻的重要性. ........................................................................................................................3
第二節 國際水稻基因定序計畫(International Rice Genome Sequencing Project, IRGSP )........................3
第三節 台灣水稻插入突變體資料庫 (Taiwan Rice Insertional Mutants Database,TRIM)...........................5
一、構建水稻T-DNA插入突變種原庫的二元載體......................................................................................6
第四節 水稻T-DNA生技研發中心(T-DNA Tagged Rice Service Center)...................................................7
第五節 T-DNA (Transfer-DNA)...................................................................................................................8
第六節 水稻基因組T-DNA插入點之分布 .................................................................................................8
第七節 插入突變法.....................................................................................................................................9
第八節 CRISPR/Cas9................................................................................................................................9
第九節 Flanking sequence 獲得之方法...................................................................................................11
一、Inverse polymerase chain reaction, IPCR.........................................................................................12
二、Adaptor ligntion mediated polymerase chain reaction, AL-PCR........................................................13
三、Thermal asymmetric interlaced-polymerase chain reaction, TAIL-PCR............................................14
四、SiteFinding-PCR, SF-PCR................................................................................................................16
五、Fusion primer and nested integrated PCR, FPNI-PCR.....................................................................18
六、Hook adaptor mediated polymerase chain reaction, HA-PCR..........................................................19
第十節 T-DNA二元載體“骨架”序列整合到水稻基因組中..........................................................................22
第三章 材料與方法...................................................................................................................................24
第一節 水稻T-DNA插入種原庫…............................................................................................................24
第二節 水稻基因組DNA之萃取. ……………………………………………………………………………….24
一、水稻樣品之清潔與處理:.....................................................................................................................24
二、陽離子界面活性劑萃取DNA法:......................................................................................................24
三、水稻基因體DNA電泳測試. ...............................................................................................................25
四、 PCR反應條件、Hooker adaptor PCR.............................................................................................27
五、PCR產物電泳分析............................................................................................................................31
六、PCR產物切膠純化步驟.....................................................................................................................31
七、定序序列與 T-DNA 相同區域之比對.................................................................................................31
八、TRIM資料庫之序列比對 (TRIM BLAST)..........................................................................................34
第三節 水稻T-DNA品系資料庫...............................................................................................................36
第四節 水稻品系T-DNA解序結果資料庫.................................................................................................38
一、提升成功率篩選之方法......................................................................................................................41
(1)解序成功...............................................................................................................................................41
(2)Backbone.............................................................................................................................................42
(3)無比對到T-DNA...................................................................................................................................43
(4)質體汙染...............................................................................................................................................43
(5)其它......................................................................................................................................................45
第五節 受菲律賓國際稻米研究中心委託解序...........................................................................................46
第六節 水稻染色體上T-DNA插入點之分布............................................................................................50
第七節 定序後比對重複序列與SNP.........................................................................................................50
第四章 結果..............................................................................................................................................56
第一節 水稻T-DNA插入種原庫...............................................................................................................56
第二節 水稻DNA萃取之結果..................................................................................................................62
第三節 水稻樣品DNA萃取後進行PCR產物統計之結果.........................................................................64
第四節 解序成功率之統計及提升成功率篩選方法....................................................................................65
第五節 受國際水稻研究中心委託難解之品系解序結….............................................................................73
第六節 水稻染色體T-DNA插入點之分布.................................................................................................75
第七節 水稻T-DNA插入點兩翼序列之重複序列結果...............................................................................77
第五章 討論..............................................................................................................................................82
第一節 水稻品系T-DNA插入種原庫........................................................................................................82
第二節 水稻DNA之萃取......................................................................................................................... 82
第三節 水稻樣品DNA萃取後進行PCR之產物統計................................................................................82
第四節 解序成功率之統計及提升成功率篩選方法....................................................................................83
第五節 受國際水稻定序中心委託難解之品系解序結果.............................................................................83
第六節 水稻基因組T-DNA插入點之分布................................................................................................85
第七節 定序後重複序列與SNP…............................................................................................................85
第七章 參考文獻..................................................................................................................................... 86
附錄..........................................................................................................................................................92
一、 種原之資料查詢及材料申請平台系統...............................................................................................92
二、 水稻突變種原庫之繁殖與入庫保存之作業流程圖.............................................................................93
附表一、目前材料及資料庫蒐集結果........................................................................................................94
附表二、受菲律賓國際稻米研究中心委託解序結果信箱內容....................................................................95






圖目錄
圖1. 國際水稻基因定序計畫各國負責解序之水稻染色體..........................................................................4
圖2. 用於基因敲除和激活標記的pTag8結構示意圖.................................................................................6
圖3. Inverse PCR原理............................................................................................................................ 12
圖4. Adaptor ligntion mediated polymerase chain reaction原理圖..........................................................13
圖5. Thermal asymmetric interlaced-polymerase chain reaction原理圖.................................................15
圖6. SiteFinding-PCR原理圖...................................................................................................................17
圖7. Fusion primer and nested integrated PCR原理圖….......................................................................18
圖8. Hooker adaptor PCR原理圖............................................................................................................20
圖9. Hooker adaptor PCR個引子位置圖.................................................................................................27
圖10. pTag8質體上T-DNA 上Right border序列及附近設計引子之位置................................................28
圖11. pTag8質體上T-DNA 上Left border序列及附近設計引子之位置...................................................28
圖12. 使用NCBI網站,將實驗室經定序後序列與T-DNA中RB序列進行比對......................................32
圖13. 送定序後與RB比對之結果...........................................................................................................32
圖14. 代號20160705-1利用TRIM進行BLAST所得知所T-DNA插入之基因體位置............................34
圖15. TRIM資料庫比對分析之結果.........................................................................................................35
圖16. 萃取水稻DNA後,將保存在-20゚C冰箱已備進行PCR反應......................................................37
圖17. PCR產物結果分成五大類…...........................................................................................................37
圖18. Hooker adaptor PCR之產物進行解序結果為Backbone................................................................42
圖19. FST經比對與質體相似度結果........................................................................................................44
圖20. QV值低於20之波型圖..................................................................................................................45
圖22. 在右邊界之已知序列上設計三條專一性引子..................................................................................47
圖23. 在T-DNA RB端設計三個引子位置................................................................................................47
圖24. 在左邊界之已知序列上設計三條專一性引子..................................................................................48
圖25. 在T-DNA RB端設計三個引子位置................................................................................................48
圖27. 品系M121741之Flanking Sequence Tag 進行比對之結果.........................................................50
圖28. 品系M121741 FST利用TRIM、NCBI blast比對結果得知插入點之位置……………...................51
圖29. 品系M121741 T-DNA 插入點於Chr12:1925958..1926337與水稻基因組中之相似度.............................................................................................................................................................53
圖30. 品系M121741 FST序列在水稻基因組中Chr12:1925958..1926337、Chr11:2006336..2006715 前後各延伸200bp之序列..............................................................................................................................................................55
圖31. 水稻T-DNA插入種原庫................................................................................................................57
圖32. 向TTRSC申請種原流程圖.......................................................................................................... 58
圖33. 利用其他水稻基因組資料庫搜尋TRIM品系..................................................................................59
圖34. T-DNA右邊界之架構......................................................................................................................65
圖35. 水稻品系序列與T-DNA RB序列進行比對..................................................................................67
圖36. 水稻樣品序列RB序列進行Blast序列比對結果............. ............. ............. .................................68
圖37. 經比對後之水稻品系之序列...........................................................................................................68
圖38. 水稻品系比對後序列與質體Backbone序列..................................................................................69
圖39. 水稻樣品上FST序列與質體上Backbone序列比對之結果............. ............. ...............................70
圖40. Hooker adaptor PCR的特性探討Backbone與T-DNA一起整合進入到水稻基因組中..............................................................................................................................................................71
圖41. PCR產物電泳探討Backbone之結果.......................................................... ................................72
圖42. 水稻在染色體上分布之相關係數..................................................................................................76
圖43. 品系M121741經左右延伸200bp 的兩條序列互相進行比對之結果. ......... ................................77
圖44. 品系M121741 CHR12:1925958..1926337、CHR11:2006336..2006715在NCBI資料庫上的插入圖及附近鄰近基因........................................ ............. ............. ............. ............. ................................79
圖45. 品系M121741 Chr11:2006336..2006715插入點與Os11g0143600放大觀察圖....................80


表目錄
表1. 各種PCR優缺點之比較.................................................................................................................21
表2. T-DNA插入水稻突變品系之Flanking sequences 擴增方法-模板PCR條件...................................29
表3. T-DNA插入水稻突變品系之Flanking sequences 擴增方法-Nest PCR反應條件..........................29
表5. T-DNA插入水稻突變品系之Flanking sequences 擴增方法-thirdly Nest PCR...............................30
表4. T-DNA插入水稻突變品系之Flanking sequences 擴增方法-Secondary Nest PCR.........................30
表6. 中央研究院余淑美教授所提供T-DNA插入突變品系之水稻葉片………………………................... 36
表7. 由 T-DNA Tagged Rice Service center(TTRSC)自2007至2016年解序成功之FST數量…………………………………………….....................………………………...........................................38
表8. 解序結果內容………….....................………………………...............................................................39
表9. 質體汙染參數設定值.....………………………..................................................................................39
表10. QV值參數設定值………………......................................................................................................40
表11. 解序成功之品系…..........................................................................................................................41
表12. Hooker adaptor PCR之產物進行解序結果為Backbone................................................................42
表13. 解序結果產物無比對到T-DNA.......................................................................................................43
表14. 解序結果產物為質體汙染..............................................................................................................43
表15. 其他未解序出T-DNA兩翼序列之原因...........................................................................................45
表16. 目前TTRSC國內外物流計有: ......................................................................................................57
表17. 目前TTRSC國內外技術移轉費用計有..........................................................................................57
表18. 各國研究室向TTRSC申請種原使用表..........................................................................................60
表19. 品系18327~184858 (代號66631~66726)DNA萃取測O.D值結果..............................................62
表20. 水稻樣品DNA萃取後進行PCR產物統計之結果..........................................................................64
表21. 利用Hooker5引子將水稻DNA進行PCR產物解序結果..............................................................65
表22. Hooker adaptor PCR之產物進行解序結果為Backbone之序列...................................................66
表23. 方法 H5 S3 S6 (RB解序)解序DNA代號1 2 3 4 5 6....................................................................73
表24. 方法 H5 H4 S3 S6 (LB解序) 解序DNA代號3 7 8 9 10..............................................................73
表25. 方法 H5 H4 S3 (LB解序)解序DNA代號 11~20..........................................................................74
表26. 方法 H5 S6 S5 S3 (RB LB解序)解序DNA代號 3 11 12 14 16 17 18.........................................74
表27. T-DNA插入水稻染色體頻率...........................................................................................................75
表28. T-DNA插入點相似度及SNP之比較..............................................................................................81
表29. Hooker adaptor PCR經感善後成本之比較....................................................................................85

許瀚升。2008。水稻T-DNA插入突變體兩翼序列自動化比對系統之建立。亞洲
大學生物資訊研究所碩士論文。
羅舜芳、余淑美。2013。台灣水稻突變種原庫與資料庫-水稻基因功能研究現況
與發展。農業生技產業季刊 36: 52-56。
鄭堯中。2007。水稻T-DNA插入突變點兩側序列解序方法之比較及插入點之研
究。亞洲大學生活應用科學研究所碩士論文。
Afolabi, A. S., Worland, B., Snape, J. W. and Vain, P. 2004. A large-scale study of
rice plants transformed with different T-DNAs provides new insight into locus composition and T-DNA linkage configurations. Theor Appl Genet . 109:815–826.
Chin, H. S., Wu, Y. P., Hour, A. l., Hong, C. Y. and Lin, Y. R. 2016. Genetic and Evolutionary Analysis of Purple Leaf Sheath in Rice. Rice a Springer Open Journal. 9: 8-22.
Cong, L., Ran, F. A., Cox, D., Lin, S., Barretto, R., Habib, N., Hsu, P. D., Wu, X. and
Jiang, W. 2013. Marraffini, L.A.; Zhang, F. Multiplex genome engineering using CRISPR/Cas systems. Science. 339: 819–823.
Cho, S. W., Kim, S., Kim, J. M., Kim, J. S. 2013. Targeted genome engineering in
human cells with the Cas9 RNA-guided endonuclease. Nat. Biotechnol. 31: 230–232.
Chang, N., Sun, C., Gao, L., Zhu, D., Xu, X., Zhu, X., Xiong, J. W. and Xi, J. J.
2013. Genome editing with RNA-guided Cas9 nuclease in zebrafish embryos. Cell Res. 23: 465–472.
Chen, C., Jin, W., Wang, M., Zhang, F., Zhou, J., Jia, Q., Wu, Y. and Wu, P. 2003.
Distribution anf characterization of over 1,000 T-DNA tags in rice genome. The Plane Journal. 36: 105-113.
Cluster, P. D., O’Dell, M., Metzlaff, M. and Flavell, R. B. 1996. Details of T-DNA
structural organization from a transgenic. Petunia population exhibiting co-suppression. Plant Mol Biol. 32: 1197–1203.
De Buck, S., Wilde, D. C., Montagu, M. V. and Depicker, A. 2000. T-DNA vector backbone sequences are frequently integrated into the genome of transgenic olants obtained by Agrobacterium-mediated transformation. Molecular Breeding. 6: 459-468.
Deng, L., Garrett, R. A., Shah, S. A., Peng, X. and She, Q. 2013. A novel interference
mechanism by a type IIIB CRISPR-Cmr module in Sulfolobus. Mol. Microbiol. 87: 1088–1099.
Dicarlo, J. E., Norville, J. E., Mali, P., Rios, X., Aach, J. and Church, G. M. 2013.
Genome engineering in Saccharomyces cerevisiae using CRISPR-Cas systems. Nucleic Acids Research. 41: 4336–4343.
Eychner, A. M., Lebo, R. J. and Elkins, K. M. 2015. Comparison of proteases in DNA extraction via quantitative polymerase chain reaction. Anal Biochemistry. 478: 128-130.
Fan, M. J., Yu, S. M., Hung, T. C., Chen, W. Q., Hsing, Y. I., Lin, H. L., Liu, C. C., Chung, G. J. and Tsai, J. J. P. 2011. TTRSIS: A Cloud Computing Platform for Rice Functional Genomics Research through a Reverse Genetics Approach. IEEE. 221-227.
Gao, S., He, D., Li, G., Zhang, Y., Lv, H. and Wang, L. 2016. A method for amplification of unknown flanking sequences based on touchdown PCR and suppression-PCR. Anal Biochemistry. 509: 79-81.
Garcia, L. T., Cristancho, L. M., Vera, E. P. and Begambre, O. 2015. A New Multiplex-PCR for Urinary Tract Pathogen Detection Using Primer Design Based on an Evolutionary Computation Method. J Microbiol Biotechnol. 25: 1714-1727.
Hsing, Y, I., Chen, C. G., Fan, M. J., Lu, P. C., Chen, K. T., Lo, S. F., Sun, P. K., Ho, S. L., Lee, K. W., Wang, Y. C., Huang, W. L, Ko, S. S., Chen, S., Chen, J. L., Chung, C. I., Lin, Y. C., Hour, A. L., Wang, Y. W., Chang, Y. C., Tsai, M. W., Lin, Y. S., Chen, Y. C., Yen, H. M., Li, C. P., Wey, C. K., Tseng, C. S., Lai, M. H., Huang, S. C., Chen, L. J. and Yu, S. M. 2007. A rice gene activation/knockout mutant resource for high throughput functional genomics. Plant Mol Biol. 63: 351-364.
Hwang, W. Y., Fu, Y., Reyon, D., Maeder, M. L., Tsai, S. Q., Sander, J. D. and Joung, J. K. 2013. Efficient genome editing in zebrafish using a CRISPR-Cas system. Nat Biotech. 31: 227-229.
Jeong, D. H., An, S., Kang, H. G, Moon, S., Han, J. J., Park, S., Lee, H. S., An, K., and An G. 2002. T-DNA insertional mutagenesis for activation tagging in rice. Plant Physiol. 130: 1636-1644.
Jeong, D. H., An, S., Park, S., Kang, H. G., Park, G. G., Kim, S. R. and An, G. 2006. Generation of a flanking sequence-tag database for activation-tagging lines in japonica rice. The Plant Journal. 45: 123-132.
Jiang, W., Bikard, D., Cox, D., Zhang, F. and Marraffini, L. A. 2013. RNA-guided editing of bacterial genomes using CRISPR-Cas systems. Nat Biotechnology. 31: 233-239.
Jinek, M., East, A., Cheng, A., Lin, S., Ma, E. and Doudna, J. 2013. RNA-
programmed genome editing in human cells. eLife. 2: e00471.
Kim, S. R., Lee, J., Jun, S. H., Park, S., Kang, H. G., Kwon, S. and An, G. 2003.
Transgene structures in T-DNA-inserted rice plants. Plant Mol Biol. 52: 761–
773.
Kononov, M. E., Bassuner, B. and Gelvin, S. B. 1997. Integration of T-DNA
binary vector “backbone” sequences into the tobacco genome: evidence for multiple complex patterns of integration. Plant J. 11: 945–957.
Lange, M., Vincze, E., Moller, M. G. and Holm, P. B. 2006. Molecular analysis of transgene and vector backbone integration into the barley genome following Agrobacterium-mediated transformation. Plant Cell Rep. 25: 815-820.
Wu, L., Di, D. W., Zhanga, D., Song, B., Luo, P. and Guoa, G. Q. 2015. Frequent problems and their resolutions by using thermal asymmetric interlaced PCR (TAIL-PCR) to clone genes in Arabidopsis T-DNA tagged mutants. Biotechnology & Biotechnological Equipment. 29: 260-267.
Lin, Y. R., Wu. Y. P., Wei, F. J., Lu, P. C., Huang, Y. C., Chang, C, H., Hour, A. L., Kou, S. C., Hsieh, J. S. and Hsing, Y. I. 2008. Construction of the Website ‘The Resource of Rice Genetic Markers in Taiwan’. Crop, Environment & Bioinformatics. 5: 1-21.
Liu, Y.G. and Chen, Y. 2007. High-efficiency thermal asymmetric interlaced PCR for amplification of unknown flanking sequences. Biotechniques. 43: 649-656.
Liu, Y. G. and Whittier, R. F. 1995. Thermal Asymmetric Interlaced PCR: Automatable Amplification and sequencing of insert end fragments form P1 and YAC clones for chromosome walking. Genomics. 25: 674-681.
Li, X. W., Wu, S. L., Liu, Y. H., Jin, G. L., Zhao, H. J., Fan, L. J. and Shu, Q.Y. 2016. Genome-wide profiling of genetic variation in Agrobacterium-transformed rice plants. Journal of Zhejiang University-SCIENCE B. 17: 992-996.
Ma, J., Wang, Y. P., Ren, S., Zhang, Z., Lu, S. and Wang, P. W. 2014. Cloning flanking sequence by single-primer PCR in transgenic plants. Genet Mol Res. 13: 8403-8410.
Maria, E. K., Bassuner, B. and Gelvin, S. B. 1997. Integration of T-DNA binary vector backbone sequences into the tobacco genome evidence for multiple complex patterns of integration. The Plant Journal. 5: 945-957.
Martineau, B., Voelker, T. A. and Sanders, R. A. 1994. On Defining T-DNA. The Plant Cell. 1032-1033.
Makarova, K. S., Aravind, L., Wolf, Y. I. and Koonin, E.V. 2011. Unification of Cas
protein families and a simple scenario for the origin and evolution of CRISPR-Cas systems. Biol. Direct. 6: 38.
Makarova, K. S., Haft, D. H., Barrangou, R., Brouns, S. J., Charpentier, E., Horvath,
P., Moineau, S., Mojica, F. J., Wolf, Y. I. and Yakunin, A. F. 2011. Evolution and classification of the CRISPR-Cas systems. Nat. Rev. Microbiol. 9: 467–477.
Marraffini, L. A. and Sontheimer, E. J. 2008. CRISPR interference limits horizontal
gene transfer in staphylococci by targeting DNA. Science. 322: 1843–1845.
Marraffini, L. A. and Sontheimer, E. J. 2010. CRISPR interference: RNA-directed
adaptive immunity in bacteria and archaea. Nat. Rev. Genetics. 11: 181–190.
Mali, P., Yang, L., Esvelt, K. M., Aach, J., Guell, M., Dicarlo, J. E., Norville, J. E.
and Church, G. M. 2013. RNA-guided human genome engineering via Cas9.
Science. 339: 823–826.
Mori, M., Tomita, C., Sugimoto, K., Hasegawa, M., Hayashi, N., Dubouzet, J.
G., Ochiai, H., Sekimoto, H., Hirochika, H. and Kikuchi, S. 2007. Isolation and molecular characterization of a Spotted leaf 18 mutant by modified activation-tagging in rice. Plant Mol. Biol. 63: 847-860.
Park, D. J., Li, R., Lau, E., Georgeson, P., Nguyen, D. T. and Pope, B. J. 2016. UNDR ROVER - a fast and accurate variant caller for targeted DNA sequencing. BMC Bioinformatics. 17: 165-172.
Qi, L. S., Larson, M. H., Gibert, L. A., Doudna, J. A., Wissman, J. S., Arkin, A. P. and
Lim, W. A. 2013. Repurposing CRISPR as an RNA-guided platform for sequence-specific control of gene expression. The Plant Cell. 152: 1173-1183.
Ramanathan, V. and Veluthambi, K. 1995. Transfer of non-T-DNA portions of the
Agrobacterium-tumefasciens Ti plasmid pTiA6 from the left terminus of T-DNA. Plant Mol Biol. 28: 1149–1154.
Sallaud, C., Gay, C., Larmande, P., Bès, M., Piffanelli, P., Piégu, B., Droc, G., Regad,
F., Bourgeois, E., Meynard, D., Périn, C., Sabau, X., Ghesquière, A., Glaszmann, J. C., Delseny, M. and Guiderdoni, E. 2004. High throughput T-DNA insertion mutagenesis in rice: a first step towards in silico reverse genetics. Plant J. 39: 450-464.
Shou, H., Frame, B. R., Whitman, S. A. and Wang, K. 2004. Assessment of
transgenic maize events produced by particle bombardment or Agrobacterium-mediated transformation. Mol Breed. 13: 201– 208.
Tan, G., Gao, Y., Shi, M., Zhang, X., He, S., Chen, Z. and An, C. 2005. SiteFinding-PCR: a simple and efficient PCR method for chromosome walking. Nucleic Acids Res. 33: e122.
Thole, V., Worland, B., Snape, J. W. and Vain, P. 2007. The pCLEAN dual binary vector system for Agrobacterium-mediated plant transformation. Plant Physiol. 145: 1211-1219.
Triglia, T., Peterson, M. G. and Kemp, D. J. 1988. A procedure for in vitro amplification of DNA segments that lie outside the boundaries of known sequences. Nucl Acids Res. 16: 8186.
Valentine, L. 2003. Agrobacterium tumefaciens and the plant: the David and Goliath of modern genetics. Plant Physiol. 133: 948-955.
Wang, D. G., Brewster, J. D., Paul, M. and Tomasula, P. M. 2015. Two methods for increased specificity and sensitivity in loop-mediated isothermal amplification. Molecules. 20: 6048-6059.
Wang, H., Fang, J., Liang, C., He, M., Li, Q. and Chu, C. 2011. Computation-assisted SiteFinding- PCR for isolating flanking sequence tags in rice. Biotechniques. 51: 421-423.
Wang, S., He, J., Cui, Z. and Shun, P. 2007. Self-formed adaptor PCR: a simple and efficient method for chromosome walking. Appl Environ Microbiol. 73: 5048-5051.
Wang, H., Yang, H., Shivalila, C. S., Dawlaty, M. M., Cheng, A. W., Zhang, F. and
Jaenisch, R. 2013. One-step generation of mice carrying mutations in multiple genes by CRISPR/cas-mediated genome engineering. Cell. 153: 910–918.
Wang, Z., Ye, S., Li, J., Zheng, B., Bao, M. and Ning, G. 2011. Fusion primer and nested integrated PCR (FPNI-PCR): a new high-efficiency strategy for rapid chromosome walking or flanking sequence cloning. BMC Biotechnol. 11: 109-121.
Wolters, A. M. A., Trindade, L. M., Jakobseb, E. and Visser, R. G. F. 1998.
Fluorescence in situ hybridisation on extended DNA fibres as a tool to analyze complex T-DNA loci in potato. Plant J. 13: 837–847.
Xu, J., Cao, J., Cao, D., Zhao, T., Huang, X., Zhang, P. and Luan, F. 2013. Flanking sequence determination and event-specific detection of genetically modified wheat B73-6-1. Acta Biochim Biophys Sin. 45: 416-421.
Yin, Z. and Wang, G. L. 2000. Evidence of multiple complex patterns of T-DNA
integration into the rice genome. Theor Appl Genet. 100: 461–470.
Zhang, J., Guo, D., Chang, Y., You, C., Li, X., Dai, X. and Wu, C. 2007. Non-random distribution of T-DNA insertions at various levels of the genome hierarchy as revealed by analyzing 13,804 T-DNA flanking sequences from an enhancer-trap mutant library. Plant J. 49: 947-959.
Zhang, S., Ravelonandro, M., Russell, P., McOwen, N., Briard, P., Bohannon, S. and Vrient, A. 2014. Rapid diagnostic detection of plum pox virus in Prunus plants by isothermal AmplifyRP(R) using reverse transcription-recombinase polymerase amplification. Journal of Virological Methods. 207: 114-120.
Zhao, G., Zhang, Z., Sun, H., Li, H. and Dai, H. 2007. Isolation of Ty1-copia-like Retrotransposon Sequences from the Apple Genome by Chromosome Walking Based on Modified SiteFinding-polymerase Chain Reaction. Acta Biochim Biophys Sin (Shanghai). 39: 675-683.
Zheng, S. J., Henken, B., Sofiari, E., jacobsen, E., Frans, A. and Kren, K. 2001. Molecular characterization of transgentic shallots (Allium cepa L.) by adaptor ligation PCR (AL-PCR) and sequencing of genomic DNA flanking T-DNA borders. Transgenic Research. 10: 237-245.

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