At2G37120: A Gene Exploration
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At2G37120: A Gene Exploration. An Arabidopsis Journey Through Experimental Questioning By Jeremy Ziskind. 112011. DNA Pool 5. What is the structure of AT2G37120?. 123257. 118631. -887 -823. -201 -180 -53 1 146 173 509 713 855 932 1212 1241.

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At2G37120: A Gene Exploration

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At2g37120 a gene exploration

At2G37120: A Gene Exploration

An Arabidopsis Journey Through Experimental Questioning

By Jeremy Ziskind


At2g37120 a gene exploration

112011

DNA Pool 5

What is the structure of AT2G37120?

123257

118631

-887 -823

-201 -180 -53 1 146 173 509 713 855 932 1212 1241

  • Expressed in Suspensor of Scarlet Runner Bean

  • Chromosome 2 of Arabidopsis

    -932 base pairs in length

    -2 exons

    -5’ and 3’ UTRs

  • What organisms is a potential ortholog found in?

    -Oryza sativa, Japanese rice

Madison Insert

SALK Inserts


What is the gene s possible function

What is the Gene’s Possible Function?

  • Codes for S1Fa which is most likely a nuclear DNA binding protein

  • S1Fa important for the binding specificity of a small protein that binds to DNA

  • Protein only has 70 amino acids (Smallest DNA binding protein ever reported)

  • Has no homology to any known DNA binding modules

  • Conclusion: S1Fa belong to a NOVEL CLASS OF DNA BINDING PROTEINS!!!


At2g37120 a gene exploration

SRB EST vs Arabidopsis

  • EST: PCSC16872 (42125) Length = 408 Score = 39.6 bits (80), Expect(3) = 2e-09 Identities = 15/21 (71%), Positives = 15/21 (71%) Frame = +2 / -2

  • Query: 353 MRNVTSRPPITSSTINPGFNP 415

  • MRNV S PP TS TI PG NP

  • Sbjct: 146 MRNVNSNPPTTSRTIKPGLNP 84

  • CONCLUSIONS: SRB EST has 71% region of amino acid homology to Arabidopsis protein sequence

Comparing AT2G37120 gene expression (protein sequence) in Arabidopsis to Scarlet Runner Bean expression


Pcl1l control

Where is S1FA mRNA present in Arabidopsis and SRB?

Conclusions: S1Fa is affected by lec1 because expression patterns are different in the absence of lec1

PcL1L Control

mRNA accumulation patterns in Scarlet Runner Bean

100 Kb Ladder

S1FA Bands

Primer Dimerazation


At2g37120 a gene exploration

Is there a knockout in Madison Lines? Rounds 1 and 2

Superpool 28

DNA Pool 5

Superpool 28

Control

-201 -180 -53 1 146 173 509 713 855 932 1212 1241

DNA Pool #5 with a size of about 1400 base pairs

Sequencing results verify that the insert is in DNA Pool #5!

DNA Pool

Screen


What is the genotype of arabidopsis plants with salk insert 118631

What is the genotype of Arabidopsis plants with SALK insert #118631?

Negative Water Control

Wild-type Allele Amplification

Mutant Allele Amplification

WT 9 8 7 6 5 4 3 2 1

Tubulin

Conclusions: SALK plants 1-9 are homozygous for the wild-type allele and have no T-DNA insertions

Extremely faint Tubulin


Further experiments

Further Experiments

  • Further SALK genotyping must be performed to determine whether At2G37120 is important for seed development

  • Phenotyping must be done on SALK #118631 plants to determine if the knockouts affect seed development or any other aspect of plant development

  • DNA Pool #5 must be narrowed down to find the individual line with a knockout in At2G37120


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