A high-throughput Arabidopsis reverse genetics system.
A collection of Arabidopsis lines with T-DNA insertions in known sites was generated to increase the efficiency of functional genomics. A high-throughput modified thermal asymmetric interlaced (TAIL)-PCR protocol was developed and used to amplify DNA fragments flanking the T-DNA left borders from approximately 100000 transformed lines. A total of 85108 TAIL-PCR products from 52964 T-DNA lines were sequenced and compared with the Arabidopsis genome to determine the positions of T-DNAs in each line. Predicted T-DNA insertion sites, when mapped, showed a bias against predicted coding sequences. Predicted insertion mutations in genes of interest can be identified using Arabidopsis Gene Index name searches or by BLAST (Basic Local Alignment Search Tool) search. Insertions can be confirmed by simple PCR assays on individual lines. Predicted insertions were confirmed in 257 of 340 lines tested (76%). This resource has been named SAIL (Syngenta Arabidopsis Insertion Library) and is available to the scientific community at www.tmri.org.
Sessions
Allen
A
Torrey Mesa Research Institute, Syngenta, 3115 Merryfield Row, San Diego, California 92121, USA. allen.sessions@syngenta.com
Burke
Ellen
E
Presting
Gernot
G
Aux
George
G
McElver
John
J
Patton
David
D
Dietrich
Bob
B
Ho
Patrick
P
Bacwaden
Johana
J
Ko
Cynthia
C
Clarke
Joseph D
JD
Cotton
David
D
Bullis
David
D
Snell
Jennifer
J
Miguel
Trini
T
Hutchison
Don
D
Kimmerly
Bill
B
Mitzel
Theresa
T
Katagiri
Fumiaki
F
Glazebrook
Jane
J
Law
Marc
M
Goff
Stephen A
SA
D016960:Agrobacterium tumefaciens / Q000235:genetics; D017360:Arabidopsis / Q000235:genetics*; D001665:Binding Sites / Q000235:genetics; D032461:Chromosomes, Plant / Q000235:genetics; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics*; D018744:DNA, Plant / Q000737:chemistry / Q000235:genetics; D030541:Databases, Genetic; D018745:Genome, Plant; D020407:Internet; D016254:Mutagenesis, Insertional; D030821:Plants, Genetically Modified; D016133:Polymerase Chain Reaction / Q000379:methods; D012639:Seeds / Q000235:genetics; D017422:Sequence Analysis, DNA
D004269:DNA, Bacterial; D018744:DNA, Plant; C036483:T-DNA