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provided <strong>the</strong> herbicide‐resistant gene BnALS1R with
independent intellectual property rights in rapeseed
breeding, as well as in o<strong>the</strong>r crops.
Cloning <strong>and</strong> characterization of
LarixgmeliniiMurEgene
Ting ZHANG 1 , Xiaofei LIN, 1 Yueying XU 1 <strong>and</strong>
Hiroyoshi TAKANO 2
1College of Life Science, Inner Mongolia University, Hohhot
010021, China <strong>and</strong> 2 Graduate School of Science <strong>and</strong> Technology,
Kumamo<strong>to</strong> University, Kumamo<strong>to</strong>, Japan.Email:
linxiaofei04@hotmail.com
The endosymbiotic <strong>the</strong>ory states that plastids of green
plants are derived from a single primary endosymbiosis
involving a eukaryote <strong>and</strong> a cyanobacterium.
Peptidoglycans (PGs) are continuous covalent
macromolecular structures found on <strong>the</strong> outside of <strong>the</strong>
cy<strong>to</strong>plasmic membrane of almost all eubacteria, which
protect bacterial cells from osmotic pressure, maintain
<strong>the</strong> unique shapes of bacterial cells <strong>and</strong> are involved in
cell division. Previous studies showed that PGs of plastids
gradually degrade during <strong>the</strong> evolution of plants. Some
genes encoded key enzyme in PGs syn<strong>the</strong>sis disappeared,
horizontally transferred or functionally changed.
Ten Mur genes are associated with PGs biosyn<strong>the</strong>sis in
bacteria. Enzymes encoded by bacterial MurE genes
catalyze <strong>the</strong> ATP‐dependent formation of uridine
diphosphate‐Nacetylmuramic acid‐tripeptide in bacterial
peptidoglycan biosyn<strong>the</strong>sis. The MurE gene have been
found in genomes of Physcomitrella patens <strong>and</strong>
Arabidopsis thaliana, <strong>and</strong> gene knockout experiments
showed that <strong>the</strong> MurE genes have different function in P.
patens <strong>and</strong> A. thaliana. PpMurE is related <strong>to</strong> chloroplast
division in moss, <strong>and</strong> AtMurE affect chloroplast
development in A. thaliana. To investigate <strong>the</strong> mechanism
of functional transformation of MurE in chloroplast
evolution, it is necessary <strong>to</strong> isolate ortholog of MurE <strong>and</strong>
analyze its function in gymnosperm.
In this study, an ortholog of MurE was isolated from Larix
gmelinii by degenerate‐PCR <strong>and</strong> RACE, referred <strong>to</strong> as
LgMurE. The 2868 bp of full‐length cDNA was predicted
<strong>to</strong> encode a protein of 787 amino acids with a molecular
mass of 86.88 kDa, which shows striking sequence
similarity <strong>to</strong> MurE proteins from bacteria, cyanobacteria
<strong>and</strong> o<strong>the</strong>r plants. A putative transit peptide predicted <strong>to</strong>
locate in chloroplast was detected from LgMurE by Target
P program. The plasmid pUC18‐TP‐GFP was constructed
<strong>to</strong> detect subcellular localization by transformation of
P.patens, which shows that <strong>the</strong> LgMurE is located in
chloroplast. To detect <strong>the</strong> function of <strong>the</strong> LgMurE,plasmid
pTFH22.4‐LgMurE sense <strong>and</strong> antisense were constructed
<strong>to</strong> introduce in<strong>to</strong> <strong>the</strong> pro<strong>to</strong>plasts of Larix gmelinii.
Dahurian larch, L. gmelinii, is an important coniferous
species, which is in a different evolutional stage with P.
patens <strong>and</strong> A. thaliana. Functional analysis of <strong>the</strong> LgMurE
<strong>and</strong> complementation assay with those of cyanobacteria,
P. patens <strong>and</strong> A. thaliana, may provide <strong>the</strong>oretical <strong>and</strong>
experimental basis for underst<strong>and</strong>ing <strong>the</strong> evolution of
plastids <strong>and</strong> <strong>the</strong> transition mechanism of gene function
during evolution his<strong>to</strong>ry.
Construction <strong>and</strong> primary analysis of
suppression subtractive library of female
sterile mutant FS­M1 in Brassica napus
Sanxiong FU, Cunkou QI, Hui GUI, Song
CHEN<strong>and</strong> Xiaoying ZHOU
Nanjing Sub‐Center (Rapeseed) of National Center of Oilseeds
Crop Improvement, Key Labora<strong>to</strong>ry of Cot<strong>to</strong>n <strong>and</strong> Rapeseed
(Nanjing), Ministry of Agriculture, China <strong>and</strong> Institute of
Industrial Crops, Jiangsu Academy of Agricultural Sciences,
Nanjing 210014, China. Email: gulemin@<strong>to</strong>ngji.edu.cn
The female sterile mutant FS‐M 1 was isolated from a
spontaneous mutation of Brassica napus var. Ningyou10.
When <strong>the</strong> mutant is crossed as a female, a very poor seed
set is obtained, whereas it is fertile as a pollen donor. The
floret of <strong>the</strong> mutant consisted of almost equal‐length
stamens, a short pistil, a flat style <strong>and</strong> ovary <strong>and</strong> <strong>the</strong>
stigma was chapped. In order <strong>to</strong> screen <strong>the</strong> genes
involved in embryogenesis <strong>and</strong> flower development, <strong>the</strong>
suppressive subtraction hybridization (SSH) technique
was used. Two differential expressing cDNA libraries
were constructed with <strong>the</strong> cDNAs of <strong>the</strong> mutant FS‐M 1 as
driver <strong>and</strong> <strong>the</strong> cDNAs of its wild type Ningyou10 as tester,
vice versa. The recombined efficiencies of <strong>the</strong> forward
<strong>and</strong> backward subtractive library were 96.2% <strong>and</strong> 94.5%,
respectively. The inserted fragment size ranged from
100bp <strong>to</strong>1000bp, with an average size of about 500 bp.
768 clones in <strong>the</strong> forward <strong>and</strong> backward subtractive
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