Non-disruptive dna isolation from corn seeds

Patent metadata

Field	Value
Publication number	US-2016115472-A1
Application number	US-201414890955-A
Country	US
Kind code	A1
Filing date	May 28, 2014
Priority date	Jun 3, 2013
Publication date	Apr 28, 2016
Grant date	—

Abstract

This invention relates to systems and process for isolating DNA from biological materials such as seeds while retaining a viable seed for further use. The seed from which the DNA is isolated remains viable and is used or discarded based on the DNA analysis of the seed soak solution. The seed soak solution can have substantially all of the confounding maternal DNA from the seed eliminated from the seed soak solutions by employing intact seed pretreatments. This method is particularly useful for maize seed.

First claim

1 . A bulked population of viable seeds, wherein substantially all of the seeds in the population have a portion of exposed seed endosperm wherein substantially all of the seeds in the population have the presence or absence of at least one targeted trait in common, and wherein the presence or absence of the at least one targeted trait in the seeds in the population is determined by analyzing the seed soaked solution from the viable exposed soak seed.

2 . The population of viable seeds according to claim 1 , wherein each seed in the population is pretreated to reduce the presence of maternal DNA from the outer portion of the seed being present in the seed soak solution.

3 . The population of viable seeds according to claim 1 , wherein each seed in the population is pretreated at least once to a soak of at least a portion of the intact seed in a DNA releasing solution to extract maternal DNA from the outer portion of the seed.

4 . The population of viable seeds according to claim 1 , wherein each seed in the population is pretreated at least once with a coating to inhibit maternal DNA from releasing out of the outer portion of the seed.

5 . The population of viable seeds according to claim 1 , wherein each seed in the population is pretreated at least once with a coating wherein said coating is a spray which can be applied paint, adhesive, glue, polymer plastics, and magnetic paint, to inhibit maternal DNA from the outer portion of the seed while maintaining seed viability.

6 . The population of viable soaked seeds according to claim 1 , wherein each seed in the population had a small portion of seed removed by an automated sampling system.

7 . The population of viable seeds according to claim 1 , wherein the at least one targeted trait comprises agronomic traits.

8 . The population of viable seeds according to claim 1 , wherein the at least one targeted trait includes a genetic marker, the genetic marker being genetically linked to at least one of a gene, a loci, an allele, and a haplotype.

9 . The population of viable seeds according to claim 1 , wherein the population comprises at least one of: at least twelve seeds, at least 500 seeds, and at least one-thousand seeds.

10 . A method for analyzing a population of seeds, the method comprising: exposing the seed endosperm, from individual seeds in a population of seeds while preserving germination viability of the seeds, soaking the viable seed in a non-disruptive solution thereby forming a seed soak solution from at least one of the individual seeds in the population of seeds and analyzing the seed soak solution for the presence or absence of one or more traits of interest.

11 . The method for analyzing a population of seeds according to claim 10 , comprising: the step of pretreating at least once, an intact seed of the population forming an intact seed solution, whereby reducing the presence of maternal DNA from the outer portion of the exposed seed in the seed soak solution.

12 . The method for analyzing a population of seeds according to claim 10 , wherein the non-disruptive solution is a non-disruptive DNA releasing or isolating solution.

13 . The method for analyzing a population of seeds according to claim 12 , wherein the non-disruptive DNA releasing or isolating solution is an alkali solution.

14 . The method for analyzing a population of seeds according to claim 10 , comprising determining the genotypic character or the phenotypic character of the seeds in the population with the seed solution isolated from the exposed viable seed.

15 . The method for analyzing a population of seeds according to claim 10 comprising: diagnosing if one or more seeds from the population exhibits the presence or absence of the one or more traits of interest; and, optionally, sorting seeds.

16 . The method for analyzing a population of seeds according to claim 10 , wherein the isolated DNA in the seed soak solution is tested for one or more traits of interest comprising a genetic marker, a single nucleotide polymorphism, a simple sequence repeat, a haplotype.

17 . The method for analyzing a population of seeds according to claim 10 , further comprising detecting diagnostically one or more seeds from the population of seeds based on the presence or absence of the one or more traits of interest.

18 . The method for analyzing a population of seeds according to claim 10 , wherein the one or more traits of interest include one or more undesirable traits of interest; the method further comprising sorting one or more seeds from the population of seeds based on the presence or absence of the one or more undesirable traits of interest, and discarding the sorted seeds.

Assignees

• Syngenta Participations Ag

Inventors

• Hannappel Ulrich Stephan

Classifications

• C12Q2600/172

  Haplotypes · CPC title

• C12Q2600/156

  Polymorphic or mutational markers · CPC title

• C12Q2600/13

  Plant traits · CPC title

• C12N15/1003 Primary

  Extracting or separating nucleic acids from biological samples, e.g. pure separation or isolation methods; Conditions, buffers or apparatuses therefor · CPC title

• C12Q1/6895

  for plants, fungi or algae · CPC title