Method for isolation of transcription termination sequences

摘要

The invention relates to efficient, high-throughput methods, systems, and DNA constructs for identification and isolation of transcription termination sequences. The invention relates further to specific terminator sequences identified by said methods isolated from rice.

主权项

1. An expression construct comprising in 5′ to 3′ direction:
a) a promoter sequence functional in plants; b) a nucleic acid sequence of interest to be expressed operably linked to the promoter sequence of a); and c) at least one sequence operably linked to the promoter sequence of a) and the nucleic acid sequence of b), wherein said at least one sequence is selected from the group consisting of:
i) the sequence of SEQ ID NO: 33;ii) sequences having a homology of at least 90% with the sequence of SEQ ID NO: 33 and capable of terminating transcription in a plant cell or organism;iii) sequences hybridizing under high stringency conditions equivalent to hybridization at 68° C. in a solution consisting of 5×SSPE, 1% SDS, 5×Denhardt's reagent and 100 μg/mL denatured salmon sperm DNA followed by washing in a solution comprising 0.1×SSPE and 0.1% SDS at 68° C. with the sequence of SEQ ID NO: 33 and capable of terminating transcription in a plant cell or organism; andiv) a fragment of at least 250 consecutive base pairs of any of the sequences of i), ii) and iii), wherein said at least one sequence of c) is heterologous to said promoter sequence of a) and/or said nucleic acid sequence of interest to be expressed of b), and is mediating termination of expression of the nucleic acid sequence of b) induced from said promoter sequence of a) at a transcription termination efficiency of at least 100.