import requests
import tensorflow as tf
import pandas as pd
import numpy as np
from operator import add
from functools import reduce
import random
import tabulate

from keras import Model
from keras import regularizers
from keras.optimizers import Adam
from keras.layers import Conv2D, BatchNormalization, ReLU, Input, Flatten, Softmax
from keras.layers import Concatenate, Activation, Dense, GlobalAveragePooling2D, Dropout
from keras.layers import AveragePooling1D, Bidirectional, LSTM, GlobalAveragePooling1D, MaxPool1D, Reshape
from keras.layers import LayerNormalization, Conv1D, MultiHeadAttention, Layer
from keras.models import load_model
from keras.callbacks import EarlyStopping, ReduceLROnPlateau
from Bio import SeqIO
from Bio.SeqRecord import SeqRecord
from Bio.SeqFeature import SeqFeature, FeatureLocation
from Bio.Seq import Seq

import cyvcf2
import parasail

import re

ntmap = {'A': (1, 0, 0, 0),
         'C': (0, 1, 0, 0),
         'G': (0, 0, 1, 0),
         'T': (0, 0, 0, 1)
         }

def get_seqcode(seq):
    return np.array(reduce(add, map(lambda c: ntmap[c], seq.upper()))).reshape((1, len(seq), -1))

class PositionalEncoding(Layer):
    def __init__(self, sequence_len=None, embedding_dim=None,**kwargs):
        super(PositionalEncoding, self).__init__()
        self.sequence_len = sequence_len
        self.embedding_dim = embedding_dim

    def call(self, x):

        position_embedding = np.array([
            [pos / np.power(10000, 2. * i / self.embedding_dim) for i in range(self.embedding_dim)]
            for pos in range(self.sequence_len)])

        position_embedding[:, 0::2] = np.sin(position_embedding[:, 0::2])  # dim 2i
        position_embedding[:, 1::2] = np.cos(position_embedding[:, 1::2])  # dim 2i+1
        position_embedding = tf.cast(position_embedding, dtype=tf.float32)

        return position_embedding+x

    def get_config(self):
        config = super().get_config().copy()
        config.update({
            'sequence_len' : self.sequence_len,
            'embedding_dim' : self.embedding_dim,
        })
        return config

def MultiHeadAttention_model(input_shape):
    input = Input(shape=input_shape)

    conv1 = Conv1D(256, 3, activation="relu")(input)
    pool1 = AveragePooling1D(2)(conv1)
    drop1 = Dropout(0.4)(pool1)

    conv2 = Conv1D(256, 3, activation="relu")(drop1)
    pool2 = AveragePooling1D(2)(conv2)
    drop2 = Dropout(0.4)(pool2)

    lstm = Bidirectional(LSTM(128,
                               dropout=0.5,
                               activation='tanh',
                               return_sequences=True,
                               kernel_regularizer=regularizers.l2(0.01)))(drop2)

    pos_embedding = PositionalEncoding(sequence_len=int(((23-3+1)/2-3+1)/2), embedding_dim=2*128)(lstm)
    atten = MultiHeadAttention(num_heads=2,
                               key_dim=64,
                               dropout=0.2,
                               kernel_regularizer=regularizers.l2(0.01))(pos_embedding, pos_embedding)

    flat = Flatten()(atten)

    dense1 = Dense(512,
                   kernel_regularizer=regularizers.l2(1e-4),
                   bias_regularizer=regularizers.l2(1e-4),
                   activation="relu")(flat)
    drop3 = Dropout(0.1)(dense1)

    dense2 = Dense(128,
                   kernel_regularizer=regularizers.l2(1e-4),
                   bias_regularizer=regularizers.l2(1e-4),
                   activation="relu")(drop3)
    drop4 = Dropout(0.1)(dense2)

    dense3 = Dense(256,
                   kernel_regularizer=regularizers.l2(1e-4),
                   bias_regularizer=regularizers.l2(1e-4),
                   activation="relu")(drop4)
    drop5 = Dropout(0.1)(dense3)

    output = Dense(1, activation="linear")(drop5)

    model = Model(inputs=[input], outputs=[output])
    return model

def fetch_ensembl_transcripts(gene_symbol):
    url = f"https://rest.ensembl.org/lookup/symbol/homo_sapiens/{gene_symbol}?expand=1;content-type=application/json"
    response = requests.get(url)
    if response.status_code == 200:
        gene_data = response.json()
        if 'Transcript' in gene_data:
            return gene_data['Transcript']
        else:
            print("No transcripts found for gene:", gene_symbol)
            return None
    else:
        print(f"Error fetching gene data from Ensembl: {response.text}")
        return None

def fetch_ensembl_sequence(transcript_id):
    url = f"https://rest.ensembl.org/sequence/id/{transcript_id}?content-type=application/json"
    response = requests.get(url)
    if response.status_code == 200:
        sequence_data = response.json()
        if 'seq' in sequence_data:
            return sequence_data['seq']
        else:
            print("No sequence found for transcript:", transcript_id)
            return None
    else:
        print(f"Error fetching sequence data from Ensembl: {response.text}")
        return None

def apply_mutation(ref_sequence, offset, ref, alt):
    """
    Apply a single mutation to the sequence.
    """
    if len(ref) == len(alt) and alt != "*":  # SNP
        mutated_seq = ref_sequence[:offset] + alt + ref_sequence[offset+len(alt):]

    elif len(ref) < len(alt):  # Insertion
        mutated_seq = ref_sequence[:offset] + alt + ref_sequence[offset+1:]

    elif len(ref) == len(alt) and alt == "*":  # Deletion
        mutated_seq = ref_sequence[:offset] + ref_sequence[offset+1:]

    elif len(ref) > len(alt) and alt != "*":  # Deletion
        mutated_seq = ref_sequence[:offset] + alt + ref_sequence[offset+len(ref):]

    elif len(ref) > len(alt) and alt == "*":  # Deletion
        mutated_seq = ref_sequence[:offset] + ref_sequence[offset+len(ref):]

    return mutated_seq

def construct_combinations(sequence, mutations):
    """
    Construct all combinations of mutations.
    mutations is a list of tuples (position, ref, [alts])
    """
    if not mutations:
        return [sequence]

    # Take the first mutation and recursively construct combinations for the rest
    first_mutation = mutations[0]
    rest_mutations = mutations[1:]
    offset, ref, alts = first_mutation

    sequences = []
    for alt in alts:
        mutated_sequence = apply_mutation(sequence, offset, ref, alt)
        sequences.extend(construct_combinations(mutated_sequence, rest_mutations))

    return sequences

def needleman_wunsch_alignment(query_seq, ref_seq):
    """
    Use Needleman-Wunsch alignment to find the maximum alignment position in ref_seq
    Use this position to represent the position of target sequence with mutations
    """
    # Needleman-Wunsch alignment
    alignment = parasail.nw_trace(query_seq, ref_seq, 10, 1, parasail.blosum62)

    # extract CIGAR object
    cigar = alignment.cigar
    cigar_string = cigar.decode.decode("utf-8")

    # record ref_pos
    ref_pos = 0

    matches = re.findall(r'(\d+)([MIDNSHP=X])', cigar_string)
    max_num_before_equal = 0
    max_equal_index = -1
    total_before_max_equal = 0

    for i, (num_str, op) in enumerate(matches):
        num = int(num_str)
        if op == '=':
            if num > max_num_before_equal:
                max_num_before_equal = num
                max_equal_index = i
    total_before_max_equal = sum(int(matches[j][0]) for j in range(max_equal_index))

    ref_pos = total_before_max_equal

    return ref_pos

def find_gRNA_with_mutation(ref_sequence, exon_chr, start, end, strand, transcript_id,
                            exon_id, gene_symbol, vcf_reader, pam="NGG", target_length=20):
    # initialization
    mutated_sequences = [ref_sequence]

    # find mutations within interested region
    mutations = vcf_reader(f"{exon_chr}:{start}-{end}")
    if mutations:
        # find mutations
        mutation_list = []
        for mutation in mutations:
            offset = mutation.POS - start
            ref = mutation.REF
            alts = mutation.ALT[:-1]
            mutation_list.append((offset, ref, alts))

        # replace reference sequence of mutation
        mutated_sequences = construct_combinations(ref_sequence, mutation_list)

    # find gRNA in ref_sequence or all mutated_sequences
    targets = []
    for seq in mutated_sequences:
        len_sequence = len(seq)
        dnatorna = {'A': 'A', 'T': 'U', 'C': 'C', 'G': 'G'}
        for i in range(len_sequence - len(pam) + 1):
            if seq[i + 1:i + 3] == pam[1:]:
                if i >= target_length:
                    target_seq = seq[i - target_length:i + 3]
                    pos = ref_sequence.find(target_seq)
                    if pos != -1:
                        is_mut = False
                        if strand == -1:
                            tar_start = end - pos - target_length - 2
                        else:
                            tar_start = start + pos
                    else:
                        is_mut = True
                        nw_pos = needleman_wunsch_alignment(target_seq, ref_sequence)
                        if strand == -1:
                            tar_start = str(end - nw_pos - target_length - 2) + '*'
                        else:
                            tar_start = str(start + nw_pos) + '*'
                    gRNA = ''.join([dnatorna[base] for base in seq[i - target_length:i]])
                    targets.append([target_seq, gRNA, exon_chr, str(strand), str(tar_start), transcript_id, exon_id, gene_symbol, is_mut])

    # filter duplicated targets
    unique_targets_set = set(tuple(element) for element in targets)
    unique_targets = [list(element) for element in unique_targets_set]

    return unique_targets

def format_prediction_output_with_mutation(targets, model_path):
    model = MultiHeadAttention_model(input_shape=(23, 4))
    model.load_weights(model_path)

    formatted_data = []

    for target in targets:
        # Encode the gRNA sequence
        encoded_seq = get_seqcode(target[0])


        # Predict on-target efficiency using the model
        prediction = float(list(model.predict(encoded_seq, verbose=0)[0])[0])
        if prediction > 100:
            prediction = 100

        # Format output
        gRNA = target[1]
        exon_chr = target[2]
        strand = target[3]
        tar_start = target[4]
        transcript_id = target[5]
        exon_id = target[6]
        gene_symbol = target[7]
        is_mut = target[8]
        formatted_data.append([gene_symbol, exon_chr, strand, tar_start, transcript_id,
                               exon_id, target[0], gRNA, prediction, is_mut])

    return formatted_data


def process_gene(gene_symbol, vcf_reader, model_path):
    transcripts = fetch_ensembl_transcripts(gene_symbol)
    results = []
    all_exons = []  # To accumulate all exons
    all_gene_sequences = []  # To accumulate all gene sequences

    if transcripts:
        for transcript in transcripts:
            Exons = transcript['Exon']
            all_exons.extend(Exons)  # Add all exons from this transcript to the list
            transcript_id = transcript['id']

            for Exon in Exons:
                exon_id = Exon['id']
                gene_sequence = fetch_ensembl_sequence(exon_id)  # Reference exon sequence
                if gene_sequence:
                    all_gene_sequences.append(gene_sequence)  # Add this gene sequence to the list
                    exon_chr = Exon['seq_region_name']
                    start = Exon['start']
                    end = Exon['end']
                    strand = Exon['strand']

                    targets = find_gRNA_with_mutation(gene_sequence, exon_chr, start, end, strand,
                                                      transcript_id, exon_id, gene_symbol, vcf_reader)
                    if targets:
                        # Predict on-target efficiency for each gRNA site including mutations
                        formatted_data = format_prediction_output_with_mutation(targets, model_path)
                        results.extend(formatted_data)
                else:
                    print(f"Failed to retrieve gene sequence for exon {exon_id}.")
    else:
        print("Failed to retrieve transcripts.")

    # Return the sorted output, combined gene sequences, and all exons
    return results, all_gene_sequences, all_exons


def create_genbank_features(data):
    features = []

    # If the input data is a DataFrame, convert it to a list of lists
    if isinstance(data, pd.DataFrame):
        formatted_data = data.values.tolist()
    elif isinstance(data, list):
        formatted_data = data
    else:
        raise TypeError("Data should be either a list or a pandas DataFrame.")

    for row in formatted_data:
        try:
            start = int(row[1])
            end = start + len(row[6])  # Calculate the end position based on the target sequence length
        except ValueError as e:
            print(f"Error converting start/end to int: {row[1]}, {row[2]} - {e}")
            continue

        strand = 1 if row[3] == '1' else -1
        location = FeatureLocation(start=start, end=end, strand=strand)
        is_mutation = 'Yes' if row[9] else 'No'
        feature = SeqFeature(location=location, type="misc_feature", qualifiers={
            'label': row[7],  # Use gRNA as the label
            'note': f"Prediction: {row[8]}, Mutation: {is_mutation}"  # Include the prediction score and mutation status
        })
        features.append(feature)

    return features

def generate_genbank_file_from_df(df, gene_sequence, gene_symbol, output_path):
    # Ensure gene_sequence is a string before creating Seq object
    if not isinstance(gene_sequence, str):
        gene_sequence = str(gene_sequence)

    features = create_genbank_features(df)

    # Now gene_sequence is guaranteed to be a string, suitable for Seq
    seq_obj = Seq(gene_sequence)
    record = SeqRecord(seq_obj, id=gene_symbol, name=gene_symbol,
                       description=f'CRISPR Cas9 predicted targets for {gene_symbol}', features=features)
    record.annotations["molecule_type"] = "DNA"
    SeqIO.write(record, output_path, "genbank")

def create_bed_file_from_df(df, output_path):
    with open(output_path, 'w') as bed_file:
        for index, row in df.iterrows():
            chrom = row["Chr"]
            start = int(row["Target Start"])
            end = start + len(row["Target"])  # Calculate the end position based on the target sequence length
            strand = '+' if row["Strand"] == '1' else '-'
            gRNA = row["gRNA"]
            score = str(row["Prediction"])
            is_mutation = 'Yes' if row["Is Mutation"] else 'No'
            # transcript_id is not typically part of the standard BED columns but added here for completeness
            transcript_id = row["Transcript"]

            # Writing only standard BED columns; additional columns can be appended as needed
            bed_file.write(f"{chrom}\t{start}\t{end}\t{gRNA}\t{score}\t{strand}\t{is_mutation}\n")

def create_csv_from_df(df, output_path):
    df.to_csv(output_path, index=False)