Xây dựng cơ sở dữ liệu SSRs từ ESTs của cây dứa

BỘ GIÁO DỤC VÀ ĐÀO TẠO
TRƢỜNG ĐẠI HỌC NÔNG LÂM TP. HỒ CHÍ MINH
BỘ MÔN CÔNG NGHỆ SINH HỌC
***000***





TRẦN NGUYỄN MINH ĐĂNG





XÂY DỰNG CƠ SỞ DỮ LIỆU SSRs (SIMPLE
SEQUENCE REPEATS) TỪ ESTs (EXPRESSED
SEQUENCE TAGS) CỦA CÂY DỨA (Ananas comosus)


/2006
BỘ GIÁO DỤC VÀ ĐÀO TẠO
TRƢỜNG ĐẠI HỌC NÔNG LÂM TP. HỒ CHÍ MINH
BỘ MÔN CÔNG NGHỆ SINH HỌC
***000***






XÂY DỰNG CƠ SỞ DỮ LIỆU SSRs (SIMPLE
SEQUENCE REPEATS) TỪ ESTs (EXPRESSED
SEQUENCE TAGS) CỦA CÂY DỨA (Ananas comosus)


Luận văn kỹ sƣ
Chuyên ngành: Công Nghệ Sinh Học



Giáo viên hƣớng dẫn: Sinh viên thực hiện:
TS. TRẦN THỊ DUNG TRẦN NGUYỄN MINH ĐĂNG
Cử Nhân LƢU PHÚC LỢI Khóa: 2002-2006


/2006
MINISTRY OF EDUCATION AND TRAINING
NONG LAM UNIVERSITY, HCMC
DEPARTMENT OF BIOTECHNOLOGY
***000***




BUILDING SSRs (SIMPLE SEQUENCE REPEATS)
DATABASE FROM ESTs (EXPRESSED SEQUENCE
TAGS) OF PINEAPPLE (Ananas comosus)



Graduation thesis
Major: Biotechnology




Professor Student
PhD. Tran Thi Dung TRAN NGUYEN MINH DANG

BSc. LƢU PHÚC LỢI Term: 2002 - 2006










Ho Chi Minh City
09/2006

iv
LỜI CẢM ƠN

XIN CHÂN THÀNH CẢM ƠN
Ban Giám Hiệu trƣờng Đại học Nông Lâm Tp. Hồ Chí Minh đã tạo mọi
điều kiện cho tôi trong suốt thời gian học tập.
Các thầy cô trong bộ môn Công Nghệ Sinh Học cùng các thầy cô đã trực
tiếp giảng dạy trong suốt bốn năm qua.
Xin bày tỏ lòng biết ơn sâu sắc đến:
 TS. Trần Thị Dung
 Cử Nhân Lƣu Phúc Lợi
Đã tận tụy hƣớng dẫn, truyền đạt kiến thức giúp cho tôi hoàn thành khóa
luận này.
Cùng toàn thể lớp Công Nghệ Sinh Học 28 thân thiện đã hỗ trợ, giúp đỡ và
động viên tôi trong suốt thời gian làm đề tài.
Thành kính ghi ơn ba mẹ cùng những ngƣời thân trong gia đình luôn tạo

điều kiện và động viên con trong suốt quá trình học tập tại trƣờng.

Tháng 08 năm 2006

Trần Nguyễn Minh Đăng

v
TÓM TẮT KHOÁ LUẬN

TRẦN NGUYỄN MINH ĐĂNG, đại học Nông Lâm TP. Hồ Chí Minh, tháng
08/2006. “XÂY DỰNG CƠ SỞ DỮ LIỆU SSRs (SIMPLE SEQUENCE
REPEATS) TỪ ESTs (EXPRESSED SEQUENCE TAGS) CỦA CÂY DỨA
(Ananas comosus)”

 
 
              





 


 Ananas comosus 


 


 
 
Ananas comosus


 


vi
MỤC LỤC

Nội dung Trang
LỜI CẢM ƠN ...............................................................................................................iv
TÓM TẮT KHOÁ LUẬN ............................................................................................. v
DANH SÁCH CÁC HÌNH ............................................................................................ x
DANH SÁCH CÁC BẢNG ..........................................................................................xi
DANH SÁCH CÁC TỪ VIẾT TẮT .......................................................................... xii
Phần 1. Mở đầu .............................................................................................................. 1
 ................................................................................................................ 1
  ........................................................................................... 1
 ..................................................................................................... 2
 ................................................................... 2
1.2. Mục tiêu của khóa luận ......................................................................................... 3
Phần 2. Tổng quan tài liệu ............................................................................................ 4
2.1. Giới thiệu về cây dứa ............................................................................................. 4
 ............................................................................................................ 4
 ............................................................................................... 4
 ..................................................................................................... 5
 ................................................................................................................. 5
 ............................................................................................................. 5

 ................................................................................................................. 5
2.1.3.4. Hoa ............................................................................................................... 5
 ............................................................................................................... 6
 ................................................................................................................ 6
 .................................................................................................... 6
2.1.4 .............................................................................................. 6
 ............................................................................................... 6
 ............................................................................................. 7
 .......................................................................................... 7
 .......................................................................................................... 8
 ............................................................................................. 8
 ................................................................................................ 9
 .............................................................................................. 9

vii
 ........................................................................................... 10
 ....................................................................... 11
 ........................................ 11
2.2. Các Marker phân tử ............................................................................................ 12
2.2.1. Isozymes .................................................................................................................... 12
2.2.2. ALP ............................................................................................................................ 12
2.2.3. AFLP .......................................................................................................................... 12
2.2.4. RAPD ......................................................................................................................... 13
2.2.5. SSCP .......................................................................................................................... 14
2.2.6. SNP ............................................................................................................................ 14
2.2.7. SSR ............................................................................................................................. 15
2.2.8.   .................................................................................. 15
2.2.9. RFLP .......................................................................................................................... 15
2.3. Chi tiết về microsatellite ..................................................................................... 16
 ................................................................................................................. 16

 ............................................................ 16
 ......................................................................................... 17
 ...................................................................................... 17
 microsatellite ........................................................................................ 18
 ................................................................................................................... 19
2.4. EST ........................................................................................................................ 19
  ......................................................................................................... 19
 ................................................................................................. 20
2.5. Cơ sở dữ liệu và hệ quản trị cơ sở dữ liệu ......................................................... 20
  ............................................................. 20
2.5.2 .......................................................... 21
2 ............................................................................ 21
  ............................................................................ 21
 ................................................................................................. 23
 .................................................................................................. 23
 ...................................................................... 23
 ............................................................................................................... 24
 .................................................... 25
 ................................................ 25

viii
 .......................................................................... 25
khai  .................................... 25
2.6. Internet và Web ................................................................................................... 26
 ................................................................................................... 26
 ........................................................................ 26
 ................................................................................. 26
 ............................................................... 28
 ............................................................................. 28
 ..................................................................................... 28

 .............................................................................. 28
2.7. Ngôn ngữ lập trình Perl và Javascript .............................................................. 28
 ........................................................................................................... 28
2.7.1.1. T ........................................................................... 28
.................................................................................................... 29
 .......................................... 29
................................................................................................. 30
 ................................................................................. 31
 ............................................................................ 31
 .............................................................. 31
2.8. Cơ sở dữ liệu sinh học ......................................................................................... 32
2.8.1. NCBI .......................................................................................................................... 32
 ......................................................................................... 32
 ............................................................... 33
 ....................................................................... 33
Phần 3. Phƣơng pháp và chƣơng trình sử dụng ....................................................... 35
3.1. Các chƣơng trình và ngôn ngữ lập trình đƣợc sử dụng .................................. 35
 ............................................................................................................. 35
  ....................................................................... 35
 .................................................... 35
 BLAST ....................... 36
 ............................................................. 36
3.1.2.4. Apache web Server .................................................................................... 37
3.2. Thu nhận trình tự SSRs ...................................................................................... 38
 ................................................................................... 40

ix
3 ............................................................................................. 41
3.3. Xây dựng CSDL, công cụ để giúp ngƣời dùng có thể khai thác tốt dữ liệu. .. 44
 ............................................................................................ 44

 ................................................................................ 44
 ............................................................................... 46
 ............................................................................... 47
3.4. Thiết kế giao diện web để truy xuất thông tin tại cơ sở dữ liệu ...................... 47
3.5. Tích hợp các công cụ sinh học vào trang web ................................................... 48
Phần 4. Kết quả và thảo luận ..................................................................................... 49
4.1. Kết quả thu nhận trình tự microsatellite .......................................................... 49
Ananas comosus.................................................... 49
 .............................................................................. 50
4.2. Xây dựng CSDL, công cụ để giúp ngƣời dùng có thể khai thác tốt dữ liệu ... 51
Ananas comosus ................................................................. 51
 ......................................... 52
 ....................................................................................................... 57
4.3. Trang web thể hiện thông tin cơ sở dữ liệu SSRs của Ananas comosus ......... 59
 ..................................................................................... 60
 .................................. 60
Ananas comosus (Ananas comosus PAGE) ......................... 60
 .............................................................. 61
 .............................................................. 62
 ........................................................................................................... 64
 .................................................. 64
Phần 5. Kết luận và đề nghị ........................................................................................ 65
5.1. Kết luận ................................................................................................................ 65
5.2. Đề nghị .................................................................................................................. 65
Phần 6. Tài liệu tham khảo ......................................................................................... 66

x
DANH SÁCH CÁC HÌNH

Hình Trang

 ...................................................... 2
 ................................................... 3
- Red Spanish  Cayenne ................................ 11
 .............................................................. 22
 ............................... 25
-DBI-DBD- ...................................... 30
 ..... 32
 ......... 38
 ................................. 39
............................. 40
 .............................. 41
........................ 42
......................................................... 42
 .................................................... 43
 ..................................................... 44
 ........................................................... 47
 .................................................. 48
 ....................................... 50
 .................................................................... 57
 ....................................................................... 59
 ................................................... 60
Ananas comosus .............................................. 61
 ESTs ............................................................................. 61
 .................................................................... 62
 ............ 63
 .................................................. 64


xi
DANH SÁCH CÁC BẢNG

Bảng Trang
 ........................................................................................ 44
 ................................................................... 45
 .......................................................................................... 45
 .................................................................................. 45
 .......................................................................................... 46
Ananas comosus  ................................................. 49
Ananas comosus  ............... 52
 ............................ 57
 ..................................................................... 51
  ............................................................... 62


xii
DANH SÁCH CÁC TỪ VIẾT TẮT

AFLP Amplified Fragment Length Polymorphism
ALP Amplicon Length Polymorphism
AP-PCR Arbitrary Primer- PCR
ASP Active Server Page
BLAST Basic Local Alignment Search Tool
CGI Common Gateway Interface
CSDL 
DBD Database Driver
DBI Database Interface
DNA deoxyribonucleic acid
EST Expressed Sequence Tag
GUI Graphical User Interface
HTML Hypertext Markup Language

HTTP Hypertext Transfer Protocol
JSP Java Server Page
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
RAPD Random Amplififed Polymorphic DNA
RDBMS Relational Database Management System
RFLP Restriction Fragment Length Polymorphism
SNP Single Nucleotide Polymorphism
SSCP Single- Strand Conformation Polymorphism
SSR Simple Sequence Repeats
STS Sequence Tagged Site

1

Phần 1
MỞ ĐẦU

1.1. Đặt vấn đề
1.1.1. Sơ lƣợc về sinh – tin học
 
.214.
 SWISS-
 

 



               

t


t
s

Nam,  , 











 
2


Hình 1. 1. Định nghĩa bioinformatics đƣợc mở rộng
1.1.2. Sơ lƣợc về cây dứa (Ananas comosus)
  

 ,  

 

  
X 
 citric, men bromelin...
 
 

1.1.3. Sơ lƣợc về phƣơng pháp Microsatellite (SSR)
ng c c li quyt vn  nh danh
v pht hiy b ng thi cng nh gi 


TOÁN HỌC
KHOA HỌC MÁY TÍNH
THỐNG KÊ
SINH HỌC
HÓA HỌC
VẬT LÝ

Bioinformatics

3


Hình 1. 2. Tìm hiểu nguồn gốc dựa vào Microsatellite
1.2. Mục tiêu của khóa luận





.

 

    Xây dựng cơ sở dữ liệu SSRs (Simple Sequence
Repeats) từ ESTs (Expressed Sequence Tags) của cây dứa (Ananas comosus)

   



Ba 
          



4

Phần 2
TỔNG QUAN TÀI LIỆU

2.1. Giới thiệu về cây dứa [9, 19]
2.1.1. Vị trí phân loại
 : Magnoliophyta
 : Liliopsida
 : Poales
 : Bromeliaceae
 : Ananas comosus (Merr.)
 : Pineapple

 : 
2.1.2. Nguồn gốc và phân bố
 
          



o

o
, kinh

o

o

(Claude, 1963)
            

 




 
 


5

n.


2.1.3. Đặc điểm hình thái
2.1.3.1. Rễ
           

 


2.1.3.2. Thân



  
 


2.1.3.3. Lá









2.1.3.4. Hoa



6






2.1.3.5. Quả
- 
     


 

2.3.1.6. Hạt




2.1.4. Đặc điểm trồng trọt
2.1.4.1. Yếu tố khí hậu


o
C  31
o

o


40
o

o
C  27
o
C.

o


o

o


2.1.4.2. Yếu tố đất đai





7





 



 




2.1.4.3. Yếu tố sinh vật
              

Dysmicocus Pseudoeocus brevipes).
Adoretus chinensis 

             

          Phytophthora
cinnamonic  Phytophthora palmisora    Erwinia
chrysanthemi

2.1.5. Giá trị kinh tế và sử dụng


        



Vitamin A.
8

Vitamin C.


Hydrat carbon: 13,7g.

Khi   
  -     
fructose.




Sau khi thu - 

              

                



2.1.6. Các giống trồng [20]
2.1.6.1. Nhóm Cayenne

 (Cayenne lisse).








9





 (Wilt).

2.1.6.2. Nhóm Queen





 Cayenne.






 
 


2.1.6.3. Nhóm Spanish (Tây Ban Nha)








10

-1,5 kg.






2.1.6.4. Nhóm Abacaxi
 









 




 
              
 

 
 
11


Hình 2. 1. Các giống dứa Natal Queen - Red Spanish – Cayenne
(
2.1.6.5. Các giống trồng trong nƣớc
 

 Na hoa (Nam phi Queen): Paris, Yellow Mauritius

 

 K
  
 
 Canning, Alexandra, Mac-
 
2.1.7. Tình hình phát triển của cây dứa trong và ngoài nƣớc
             


 




 
.

284.
12

2.2. Các Marker phân tử [2, 4]
2.2.1. Isozymes
Isozymes             

              



e:


2.2.2. ALP (Amplicon Length Polymorphism)



2.2.3. AFLP (Amplified Fragment Length Polymorphism)
 
 AFLP: k
DNA DNA DNA 

 

.
DNA.
:



 

 .
.
13

2.2.4. RAPD (Random Amplififed Polymorphic DNA)







  
             

 


ra AP-PCR (Arbitrary Primer- PCR).
Trong AP-
             

 
 

0





 -
t nhau.



 
