BigDNA: Primer Design Software for Overlap-Based Assembly of Phage Genomes and Larger DNAs.

PHAGE (New Rochelle, N.Y.) Pub Date : 2022-12-01 Epub Date: 2022-12-19 DOI:10.1089/phage.2022.0033

Ivan Vuong, Catherine M Mageeney, Kelly P Williams

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Abstract

Background: Gibson assembly and assembly-in-yeast are strategies to create long synthetic DNAs from diverse fragments, for example, when engineering bacteriophage genomes. Design for these methods requires terminal sequence overlaps in the fragments, determining the order of assembly. Design to rebuild a genomic fragment that is too long for a single PCR presents a puzzle since some candidate joint regions cannot yield satisfactory primers for the overlap. No existing overlap assembly design software is open-source, and none explicitly supports rebuilding.

Methods: We describe here bigDNA software that solves the rebuilding puzzle by recursive backtracking, with options to remove or introduce genes; it also tests for mispriming on the template DNA. BigDNA was tested with 3082 prophages and other genomic islands (GIs), from 20 to 100 kb, and the synthetic Mycoplasma genitalium genome.

Results: Rebuilding assembly design succeeded for all but ∼1% of GIs.

Conclusion: BigDNA will speed and standardize assembly design.

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BigDNA:噬菌体基因组和较大dna重叠组装的引物设计软件。

背景:Gibson组装和酵母内组装是一种从不同片段中合成长dna的策略，例如在噬菌体基因组工程中。这些方法的设计要求片段的末端序列重叠，决定了组装的顺序。设计重建一个对于单个PCR来说太长的基因组片段是一个难题，因为一些候选的连接区域不能产生令人满意的重叠引物。没有现有的重叠装配设计软件是开源的，也没有明确支持重建。方法:我们在这里描述了bigDNA软件，它通过递归回溯解决了重建难题，具有删除或引入基因的选项;它还能检测模板DNA上的引物错误。BigDNA在3082个原噬菌体和其他基因组岛(gi)上进行了测试，这些基因组岛的长度从20到100 kb不等，BigDNA还与合成的生殖支原体基因组进行了测试。结果:除了1%的GIs外，所有GIs的重建装配设计都成功了。结论:BigDNA将加快和规范装配设计。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

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PHAGE (New Rochelle, N.Y.)

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