The Development of Chromosome Microdissection and Microcloning Technique and its Applications in Genomic Research

Zhou, Ruonan; Hu, Zhongliang

doi:10.2174/138920207780076929

Cited by 39 publications

(25 citation statements)

References 61 publications

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“…This approach might be challenging to apply for very young sex chromosomes that might have not diverged enough in size and GCcontent from each other. However, individual chromosomes can also be enriched by microdissection (Zhou and Hu 2007) or laser capture dissection (Keinath et al 2015), and the computational techniques developed here also have the potential to be utilized in such situations, thus increasing the applicability of the method. Therefore, our approach provides a timely opportunity to generate data needed for the studies on sex chromosome evolution and sexbias in dispersal across populations.…”

Section: Discussionmentioning

confidence: 99%

A time- and cost-effective strategy to sequence mammalian Y Chromosomes: an application to the de novo assembly of gorilla Y

et al. 2016

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The mammalian Y Chromosome sequence, critical for studying male fertility and dispersal, is enriched in repeats and palindromes, and thus, is the most difficult component of the genome to assemble. Previously, expensive and labor-intensive BAC-based techniques were used to sequence the Y for a handful of mammalian species. Here, we present a much faster and more affordable strategy for sequencing and assembling mammalian Y Chromosomes of sufficient quality for most comparative genomics analyses and for conservation genetics applications. The strategy combines flow sorting, short-and long-read genome and transcriptome sequencing, and droplet digital PCR with novel and existing computational methods. It can be used to reconstruct sex chromosomes in a heterogametic sex of any species. We applied our strategy to produce a draft of the gorilla Y sequence. The resulting assembly allowed us to refine gene content, evaluate copy number of ampliconic gene families, locate species-specific palindromes, examine the repetitive element content, and produce sequence alignments with human and chimpanzee Y Chromosomes. Our results inform the evolution of the hominine (human, chimpanzee, and gorilla) Y Chromosomes. Surprisingly, we found the gorilla Y Chromosome to be similar to the human Y Chromosome, but not to the chimpanzee Y Chromosome. Moreover, we have utilized the assembled gorilla Y Chromosome sequence to design genetic markers for studying the male-specific dispersal of this endangered species.

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Section: Discussionmentioning

confidence: 99%

A time- and cost-effective strategy to sequence mammalian Y Chromosomes: an application to the de novo assembly of gorilla Y

et al. 2016

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“…Chromosomal microdissection followed by FISH (fluorescent in situ hybridization) represents a powerful tool to locate target DNA sequences allowing evaluation of the whole genome, chromosomes or even subchromosomal regions depending on the complexity of probes (Zhou and Hu 2007).…”

Section: Introductionmentioning

confidence: 99%

Similarity of heterochromatic regions in the stingless bees (Hymenoptera: Meliponini) revealed by chromosome painting

et al. 2014

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Most Meliponini share a distinctive pattern of heterochromatin distribution in relation to other bees. In general, they present one euchromatic and one heterochromatic chromosome arm, a feature explained by minimum interaction theory, which involved centric fissions followed by heterochromatin addition. In this work, two Meliponini with a distinct pattern of heterochromatin distribution, Tetragonisca fiebrigi and Melipona rufiventris, were analyzed using chromosomal microdissection of the heterochromatin region followed by FISH (fluorescent in situ hybridization). Hybridization revealed FISH signals equivalent to location of the isolated fragment that were widespread over heterochromatic portions of other chromosomes. This result showed that the heterochromatic sequences were very similar among chromosomes in the same species. Cross-hybridization of each probe in M. rufiventris and T. fiebrigi yielded no signals, revealing that both species presented differentiated and non-homologous heterochromatin sequences.

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“…Techniques such as microdissection and fluorescent in situ hybridization (FISH) has especially contributed to cytogenetic studies, making them new alternatives for research on the genome structure of organisms. This methodology eventually established a connection between classical cytogenetics and molecular genetics allowing for a series of applications, such as the production of specific probes, chromosome painting and physical genome mapping (Zhou and Hu 2007), including the visualization of rearrangements and polymorphisms not detected by conventional cytogenetics (Guerra 2004). This modality of study offers a greater resolution in chromosome analyses and enables the location of target sites in any portion of the karyotype.…”

Section: Introductionmentioning

confidence: 99%

Microdissection: a tool for bee chromosome studies

et al. 2011

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-The emergence of new molecular biology techniques has provided cytogenetics with tools which allow for the elucidation of questions that classical cytogenetics could not answer. Therefore, the present work standardizes a microdissection protocol for cytogenetic studies in bees. This methodology was first used in these insects and may contribute greatly to studies involving chromosomal rearrangements, heterochromatin composition, B chromosomes and others. For this study, the centromeric region of chromosomes in the stingless bee Tetragonisca fiebrigi was used for probe synthesis. The results demonstrated that the methodology used was efficient, presenting markings in the centromeric regions of several chromosomes. Hybridization in other sites indicates that the probe was able to detect regions that present homology with its sequence. This indicates that the technique is effective to study chromosomal evolution, genome organization and even the origin of B chromosomes.

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The Development of Chromosome Microdissection and Microcloning Technique and its Applications in Genomic Research

Cited by 39 publications

References 61 publications

A time- and cost-effective strategy to sequence mammalian Y Chromosomes: an application to the de novo assembly of gorilla Y

A time- and cost-effective strategy to sequence mammalian Y Chromosomes: an application to the de novo assembly of gorilla Y

Similarity of heterochromatic regions in the stingless bees (Hymenoptera: Meliponini) revealed by chromosome painting

Microdissection: a tool for bee chromosome studies

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