An optimal pooling strategy applied to high-throughput screening for rare marker-free transformants.

Abstract:

:An optimal pooling system, called Accurate and Fast Target Screening, has been developed for high-throughput identifying the rare marker-free transformants. This system can identify targets between 10- and 100-fold more efficiently than analysis of individual samples. By calculating the efficiency for different proportions of targets and the optimal group size in a worst case scenario, we are able to estimate an upper limit for the number of tests that are required. The application of this system to determine the transgene in an artificially constructed population of transgenic and non-transgenic wheat lines successfully identified the 10 positive samples located randomly with 990 negative samples using only 92 PCR reactions. The same approach was also applied to determine transgene expression by SDS-PAGE of seed proteins. This system gives unambiguous positive or negative results and should facilitate marker-free transformation.

journal_name

Biotechnol Lett

journal_title

Biotechnology letters

authors

Zhang J,Xiao Q,Li K,Chen M,Chang J,Luo L,Li Y,Liu Y,Shewry PR,He G

doi

10.1007/s10529-006-9129-0

subject

Has Abstract

pub_date

2006-10-01 00:00:00

pages

1537-44

issue

19

eissn

0141-5492

issn

1573-6776

journal_volume

28

pub_type

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