Karyotype analysis is one of the main techniques of cytogenetics through medical image processing, which has an important role in modern medical treatment and diagnosis. The process of human karyotype analysis contains two key components: Firstly, chromosomes are segmented from metaphase chromosome digital images taken under a microscope. Chromosomes then are analyzed, compared, ordered and classified one by one carefully. Under this procedure, the operation on segmentation and classification is cumbersomely time-consuming, where traditional geometric or statistical methods only have limited effect due to low accuracy. Thus, in most conditions, human effort is still heavily required to monitor the workflow and correct the errors. In this paper, we present an integrated workflow to segment out and classify chromosomes automatically using a combination of multiple input convolutional neural networks (CNN) and geometric optimization, called mCNN_GO. We investigate Mask R-CNN to segment out chromosomes from metaphase chromosome images and train the mCNN_GO to classify the sub-images. To improve the performance of the segmentation network, we adapt a new feature-based approach to synthesize images on the labeled data. Furthermore, we develop a geometric algorithm to straighten the chromosomes before classification to ensure the consistency of the data. Experimental results demonstrate that our approach significantly outperforms the state-of-the-art methods on automatic karyotype analysis. INDEX TERMS Biomedical image processing, image processing, machine learning.