Are At3G60030 and At2G47070 Lethal to Seed Development? - PowerPoint PPT Presentation

Title: Are At3G60030 and At2G47070 Lethal to Seed Development?


1
Are At3G60030 and At2G47070 Lethal to Seed
Development?
Jason Chai HC70AL
2
What are my genes?
At3G60030
At2G47070
3
Relevant Information of Genes

• At3G60030
• 3rd chromosome
• Opposite orientation
• - 3717 bp long
• 928 amino acids
• 10 exons
• 9 introns

• At2G47070
• 2nd chromosome
• 5 ? 3 orientation
• 4173 bp long
• 882 amino acids
• 10 exons
• 9 introns

4
What Protein Does My Genes Encode?

• At3G60030
• Squamosa promoter-binding protein
• SPL 12
• Plant-specific transcription factor
• Related to flower development

• At2G47070
• Squamosa promoter-binding protein
• SPL 1
• Plant-specific transcription factor
• Related to flower development

5
Important Information of Protein

• The Arabidopsis thaliana Squamosa binding
  protein-like (SPL) gene family represents a group
  of structurally diverse genes encoding putative
  transcription factors found only in plants.
• The distinguishing characteristic is the SBP-box
  encoding a highly conserved protein domain of 80
  amino acids in length.
• This SBP-domain is responsible for the
  interaction with DNA.
• SBP-box genes play a role in plant development,
  mainly in the flowers.

6
SPL Family

• SPL8 mutation
• Affects pollen sac development in Arabidopsis.
• Anther at stage 9 to 10
• Mutant only bears two pollen sacs with few
  microspores.
• Wildtype contains more pollen sacs with
  free-floating microspores.

Figure 4. Light Microscopic Analysis of
Developing spl8 Mutant Anthers. (A) Cross-section
through spl8-1 mutant anthers comparable to stage
4 to 5 of wild-type anther development. spl8-1
mutant anthers often develop fewer than four
pollen sacs (arrowheads). (B) An spl8-1 mutant
pollen sac at a higher magnification to
illustrate the often-observed disturbed tissue
differentiation. In particular, the outer
secondary parietal cells may not undergo a
further mitotic division and remain
undifferentiated. Intermittently, in the
developing outer secondary parietal layer,
formation of presumptive endothecium and middle
layer cells caused by periclinal divisions
(arrows) can be seen. Furthermore, the cells of
the inner secondary parietal layer or tapetum are
less well differentiated, reduced in number, and
have an appearance closer to that of the
sporogenous cells. Sporogenous cell number also
is reduced compared with that in the wild
type. (C) The failure of spl8-1 sporogenous cells
to undergo meiosis, as observed after stage 5 of
wild-type anther development, and the subsequent
decay of the pollen sac is revealed by the dense
staining of their cytoplasm and the vacuolated
tapetal cells. (D) Somewhat oblique cross-section
through a wild-type pollen sac at stage 5 to 6 of
anther development just before meiosis. All cell
layers are well differentiated, and dark-staining
callose surrounds the meiocytes. (E)
Cross-section through an spl8-1 mutant anther
comparable to stage 9 to 10 of wild-type anther
development (cf. F). This anther bears two
pollen sacs with some free microspores. (F)
Cross-section through a wild-type anther at stage
9 to 10. All pollen sacs are well developed and
contain free vacuolated microspores with exine
walls. ct, connective tissue en, endothecium
ep, epidermis g, gynoecium mc, meiocyte ml,
middle layer msp, microspore osp, outer
secondary parietal cell layer p, petal s,
sepal sc, sporogenous cells ta, tapetum vc,
vascular cells. (B) to (D) are at the same scale,
as are (E) and (F). Bars 25 µm in (A), 10 µm in
(C), and 50 µm in (F).
7
What are the genotypes of Plant 1?
Expected size of WT 1841 bp Total
Homozygous mutant 7 Expected size of Mutant 620
bp Total Wildtype 5 Total
Heterozygous 4
8
What are the Genotypes of Plant 2?
Plant 23
Plant 10
Plant 12
Plant 13
Plant 14
Plant 11
Plant 15
Plant 16
Plant 17
Plant 18
Plant 19
Plant 20
Plant 21
Plant 22
Plant 24
Wildtype
Positive
Negative
Expected size of WT 1514 bp
Total Wildtype 24
9
Where is At3G60030 active?
Expected size of RT-PCR 452 bp
10
Where is At2G47070 active?
Expected size of RT-PCR 369 bp
11
Are there any phenotypical differences?
Wildtype
vs At3G60030
12
Cloning the Upstream of At3G60030
13
Cloning the Upstream of At2G47070
14
Special Thanks to the Officers in Charge

• General Dr. Bob Goldberg
• Lieutenant General Dr. Ahnthu Bui
• Major General Dr. Xingjun Wang
• Brigadier Generals Tomokazu Kawashima Brandon
  Le
• Colonels Mike Gavino, Jonathan Russell, Ria
  Yagnik Jessica Luke
• Privates Melissa, Thi, Matt, Jordan, Jennifer,
  Bekah, Brian, Jason, Daisy, Heather