i
BỘ GIÁO DỤC VÀ ĐÀO TẠO
TRƢỜNG ĐẠI HỌC NÔNG LÂM TP. HỒ CHÍ MINH
BỘ MÔN CÔNG NGHỆ SINH HỌC
************ KHÓA LUẬN TỐT NGHIỆP
KHAI THÁC DỮ LIỆU ESTs (EXPRESSED SEQUENCE
TAGs) Ở CHI CAM CHANH (CITRUS) CHO VIỆC PHÁT
TRIỂN MARKER PHÂN TỬ SSR (SIMPLE SEQUENCE
REPEATS) Ngành học: CÔNG NGHỆ SINH HỌC
Niên khóa: 2003-2007
Sinh viên thực hiện: LƢU TRẦN CÔNG HUY Thnh ph H Ch Minh
Thng 9/2007
iii
TÓM TẮT KHOÁ LUẬN
LƢU TRẦN CÔNG HUY, Đại Học Nông Lâm TP. Hồ Chí Minh, tháng
07/2007. “KHAI THÁC DỮ LIỆU ESTs (EXPRESSED SEQUENCE TAGs) Ở
CHI CAM CHANH (CITRUS) CHO VIỆC PHÁT TRIỂN MARKER PHÂN
TỬ SSR (SIMPLE SEQUENCE REPEATS)”



 



                
xpressed Sequence Tags), trong
citrus)
Simple Sequence Repeats




sau:



3 v
ABSTRACT
LUU TRAN CONG HUY, NONG LAM UNIVERSITY, DATA MINING
FOR DEVELOPING SIMPLE SEQUENCE REPEATS (SSR) MARKER IN
EXPRESSED SEQUENCE TAGS (ESTs) FROM CITRUS
Supervisor:


The research was carried out at the department of biotechnology at Nong
Lam University.
Recent advances in genomic technologies have generated a vast amount of
publicly available expressed sequence tags (ESTs) in Citrus. These data can be
mined to identify Simple sequence repeats (SSRs) or microsatellites. These SSRs
are useful because of a broad range of application, such as genome mapping and
characterization, phenotype mapping, marker assisted selection of plant breeding,
additional map-based cloning of important genes. Moreover, this method of
developing SSR marker from ESTs is inexpensive comparing to the traditional
methods.
Methodology
1) We used perl script to receive EST sequences from database NCBI
2) Finded and separated SSRs include in ESTs database
3) We were learning about relationship database model to used to saved
vii
Mục Lục
 .................................................................................................... iii
 ................................................................................. iv
ABSTRACT ...................................................................................................... vi
 ................................................................ xi

Chƣơng 1 ............................................................................................................ 1
 ............................................................................................................. 1
1.1 Đặt vấn đề
1.2.Mục tiêu của khóa luận

Chƣơng 2 ............................................................................................................ 3
 ................................................................................... 3
 ........................................................................... 3
 ........................................................................................... 3

 BLAST .................. 22
 ..................................... 23
3.1.2.4 Egassembler .......................................................................................... 23
3.1.3 Apache web Server .................................................................................. 24
 ......................................................................... 25

Chƣơng 4 .......................................................................................................... 37
 .......................................................................... 37
4.1  ....................................... 37
EGassembler 
........................................................................................................................... 38
 ..................................................................................... 38
4.2.2  ........................... 39
 .................................... 39

ix
4.3 Assembling .................................................................................................. 41
  .............................. 42
4.4.1 BLASTn: ................................................................................................. 43
4.5. ......................................................................... 45
4.6 tBLASTx ..................................................................................................... 48
4
 ............................................................................................ 49
        
 .......................................................................................... 49
 ...................................................................... 49
SRs (SSRs PAGE) ................................................. 50

Chƣơng5 ........................................................................................................... 52
 ............................................................................... 52

DBI Database Interface
DNA deoxyribonucleic acid
EST Expressed Sequence Tag
HTML Hypertext Markup Language
HTTP Hypertext Transfer Protocol
NCBI the National Center for Biotechnology Information
NIG the National Institute of Genetics
NIH the National Institutes of Health
NLM the Nation Library of Medicine
Perl Practical Extraction and Report Language
PHP Hypertext Preprocessior
RDBMS Relational Database Management System
SNP Single Nucleotide Polymorphism
SSCP Single- Strand Conformation Polymorphism
SSR Simple Sequence Repeats
STS Sequence Tagged Site
xi
DANH SÁCH CÁC BẢNG

 .................. 26
  ............................ 26

 ............................................................................ 8
 ................................................. 12
 ............................................. 13
 .......................... 16
 ...... 19

www.NCBI.nlm.nih.gov/genomes/plant/plantlist.html#est) ............................ 27
2 Egassembler .............................................. 29
 .................................................... 30
 ............................................... 31
 ........................................... 31
 ......................................... 32
         -
india.org/ssr/ssr.htm) ......................................................................................... 36
 .......................................... 37
 ................... 40
 ............................................... 41
 ... 42-43
 ............ 44
 .................... 46
 .............. 47
 .............................................................. 49
 ...................................................................... 49
 ........................................................ 50

 ...................................................................................... 51
Chƣơng 1
MỞ ĐẦU
1.1 Đặt vấn đề
   

3. 

4. K-
Egassembler)
5. 

6. 

7. 
 3
Chƣơng 2
TỔNG QUAN TÀI LIỆU
2.1 Giới thiệu về chi cam chanh
Chi Cam chanh (Citrus)  trong 
(Rutaceae).
  -15 m

 -
 

4--
 


2.1.1 Vị trí phân lọai
 Plantae
 Magnoliophyta
Citrus sinensis x Poncirus trifoliata Citrus aurantium
5
Citrus Unshiu


2.1.3 Sâu hại và bệnh tật
Bệnh do virus
Virus citrus 

 


             
vir


Virus Tristeza (CTV)
Tristeza 

Tristeza 
Virus Tristeza   




 


Hình 2.1. CTV dƣới KHV điện tử

8


Hình 2.2: Nguồn gốc của EST
2.3.Sơ lƣợc về phƣơng pháp Microsatellite (SSR)
2.3.1Những khái niệm về kỹ thuật microsatellite
Micro         
(q.v.). M

 

 





microsatellite                

nucleotide CA  



10



    

- s


 




              


2.3.2.2 Khuếch đại của microsatellites


                  

 








12
  
 





2.3.3 Các loại microsatellite
-
Dinucleotide SSR (GT)6
GTGTGTGTGTGT
Trinucleotide SSR (CTG)4
CTGCTGCTGCTG
Tetranucleotide SSR (ACTC)4
ACTCACTCACTCACTC
           
., 1996).
2.3.4 Cơ chế hình thành microsatellite



Quá trình bắt chéo lỗi trong quá trình giảm phân (unequal crossing- over
during meiosis)
.