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Karyotype analysis of metaphase chromosomes for clinical diagnosis can be done manually or using automated software. Automated karyotyping software provides a reliable, accurate, fast, and cost effective method to analyze the whole genome of a patient. Abundant automatic karyotyping software has been developed and tested over the past 40 years [ 16 ]. The most important features and applications of existing automatic karyotyping software have been summarized as follows [ 20 ].
Agena®, InForm, and Fraenkel Systeme are the main suppliers of automated karyotyping equipment. Though their products are similar in terms of hardware and software, they are quite different in software algorithms. The software algorithms of Agena, InForm, and Fraenkel Systeme are almost identical and are further described below. Agena System is based on the previously published software developed by Fraenkel [ 21 ]. The Agena system is available in two software versions: MSK and NME. The software version used in this study was MSK software version 9.0.2. Inform software is based on the Fraenkel and KaryoWorks software [ 22 ], and was further modified and developed by the authors of this article and collaborators at the University of Texas at Austin, Baylor College of Medicine, and Lawrence Livermore National Laboratory. A soft-dependent calculation was modified in the workflow in order to increase the accuracy. The software version used in this study was: KaryoWorks v9.2. Numerical Ploidy is a free and open-source software package that is widely used for chromosomes analysis in clinical diagnosis. It has a very powerful and rich graphical interface with several preset algorithms and complete automation [ 23 ]. Numerical ploidy is based on the original software designed by Rieker et al [ 23 ].
The karyotypes of the 70 cases using CMA, karyotype analysis, and cytogenetic G-banding were considered normal. There were no discrepancies between the CMA results and the results obtained with cytogenetic G-banding analysis.
Twelve cases in the normal group were found to have benign clinical CNVs using whole genome sequencing and whole exome sequencing, but these variants were absent on cytogenetic G-banding analysis and CMA, which showed normal karyotype status (non-pathologic variants). There were 45 clinically significant pathogenic CNVs (pCNVs) in 21 cases of VOUS, and 3 clinically significant pCNVs were found in chromosomes and chromosomes including abnormal karyotypes: female, 15; male, 1. Furthermore, 15 CNVs were present in chromosomes that also had abnormal karyotypes; 11 were present in chromosomes with normal karyotypes (938.55%, 15/17). These results indicate that VOUS CNVs that do not show an association with abnormal karyotypes should be further evaluated using other methods (e.g., whole genome sequencing and exome sequencing) to identify other associated abnormalities, to elucidate the pathogenicity of the detected variants, and to determine whether there is a causal relationship between the pCNV and the abnormal karyotype [ 24 ].
Metaheuristic search-based techniques have also been widely used for the evolutionary engineering of synthetic chromosomes. The interested reader is referred to metaheuristics. In this section, we use a backtracking recombination algorithm [ 32 ] and generation of diploid chromosome sets to analyze karyotype data.
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